SESSION IV - PLANT ARCHITECTURE

ORAL COMMUNICATION

Proceedings of the XLVII Italian Society of Agricultural Genetics - SIGA Annual Congress
Verona, Italy - 24/27 September, 2003
ISBN 88-900622-4-X

Oral Communication Abstract - S4a

REGULATION OF PHYLLOTAXIS VIA POLAR AUXIN TRANSPORT

C. KUHLEMEIER 

Institute of Plant Sciences, University of Berne, CH-3013 Berne, Switzerland

Leaves and flowers are formed at the shoot apical meristem according to specific patterns, called phyllotaxis. The beauty and regularity of phyllotaxis have for centuries attracted the attention of philosophers, mathematicians and natural scientists, but experimental approaches have been few. Previous studies have shown that the plant hormone auxin triggers organ formation. Auxin undergoes specific polar transport, via cellular influx and efflux carrier proteins. I will discuss our experiments that show how these auxin carriers regulate phyllotaxis. Our data indicate that auxin is acropetally transported into the meristem through the epidermis and the meristem L1 layer. In the meristem, auxin becomes redistributed by the primordia which function as sinks for auxin. Hence, the pre-existing primordia dictate the position of future primordia by allowing auxin accumulation in the meristem only at certain minimal distances. The emerging model for phyllotaxis accounts for its reiterative nature, as well as for its regularity and stability.

Proceedings of the XLVII Italian Society of Agricultural Genetics - SIGA Annual Congress
Verona, Italy - 24/27 September, 2003
ISBN 88-900622-4-X

Oral Communication Abstract - S4b

KNOX GENES IN PLANT HORMONE METABOLISM AND SIGNAL TRANSDUCTION

G. FRUGIS*, D. MARIOTTI*, N.-H. CHUA**

*) Istituto di Biologia e Biotecnologia Agraria (IBBA), CNR, Area della Ricerca di Roma, Via Salaria Km. 29,300, 00016 Monterotondo Scalo (Roma), IT
**) Laboratory of Plant Molecular Biology, Rockefeller University, 1260 York Avenue, 10021 New York, NY – USA

hormones, transcription factors, Arabidopsis thaliana, KNOX

KNOX transcription factors have an important role in the shoot apical meristem formation a maintenance. Overexpression of knox genes, similar to the KN1 gene from maize, in different species causes strong alteration of cell fate with dramatic changes of the whole plant architecture and leaf morphology. Several studies have suggested that knox genes might affect cytokinins and gibberellins homeostasis as transgenic plants or mutants that misexpress knox factors display altered levels of these hormones. 

STM is the only KNOX gene which mutation completely prevents embryonic shoot apical meristem formation. Weak stm alleles display alteration of meristem activity and abnormal flower morphology. On this base, it was hypothesized that STM is required for both SAM formation and function during the whole life of the plant. STM target genes have not been isolated  so far and little information is available on the possible hormone pathways through which it may act to control meristem formation and activity during different phases of plant development. Evidences for genetic interactions among STM, CUC genes (member of the NAC transcription factor family) and PIN1 and MP (genes involved in auxin transport and localization) have been recently found.

To better characterize STM function in plant development and a possible connection to auxin transport, we produced and characterize Arabidopsis transgenic plants in which STM expression is either increased (constitutively or in a glucocorticoid-induced fashion) or silenced through RNA interference technology. We also analyzed auxin distribution and both auxin and gibberellin responses in plants that misexpress STM in order to identify the hormone biosynthesis or signaling pathways through which STM exerts its key role in meristem function.

Proceedings of the XLVII Italian Society of Agricultural Genetics - SIGA Annual Congress
Verona, Italy - 24/27 September, 2003
ISBN 88-900622-4-X

Oral Communication Abstract - S4c

DOWNREGULATION OF AN EXPANSIN GENE MODULATES THE PROTEIN TO CELLULOSE RATIO IN CELLWALLS AND LEADS TO PHENOTYPIC CHANGES IN PETAL LIMB

S. ZENONI*, L. REALE**, G. B. TORNIELLI*, L. LANFALONI***, A. PORCEDDU****, A. FERRARINI*, A. ZAMBONI*, C. MORETTI**, F. FERRANTI**, M. PEZZOTTI*

*) Dipartimento Scientifico e Tecnologico, Università degli Studi di Verona
**) Dipartimento di Biologia Vegetale e Biotecnologie Agroambientali, Università degli Studi di Perugia
***) Dipartimento di Biologia Cellulare e Molecolare, Università degli Studi di Perugia
****) Istituto di Genetica Vegetale- CNR

cell expansion, cellulose, downregulation, expansin, Petunia hybrida

In plants, organ shape is determined by organised and regulated control of cell expansion together with cell division. Emerging data support the view that the direction and magnitude of the enlargement of the primary cell wall largely determine the expansion pattern and thereby the final shape and size of the cells. 

Cell wall proteins are believed to play important roles in regulating cell wall extensibility which is a key parameter determining cell expansion. Among cell wall proteins studied to date, expansins are unique in the ability to induce immediate cell wall extension in vitro and cell expansion in vivo. Expansin genes have been identified in many plant species and make up a large superfamily that is divided into two major families, alpha- and beta-expansins, on the basis of the sequence divergence and biochemical activity. During flower development, floral organs typically assume their final shape after mitotic activity has basically ceased, indicating that cell expansion plays an important role in determining organ shape. 

To initiate the investigation on the function of expansins in vivo and during plant development, we cloned PhEXP18, a Petunia hybrida alpha-expansin gene, determined its pattern of spatial and temporal expression in plant and flower organs and generated transgenic Petunia lines that express the cDNA encoding PhEXP18 in the antisense orientation under control of the strong and constitutively expressed cauliflower mosaic virus (CaMV) 35S promoter. PhEXP18 was expressed mainly in petals, ovaries and stems. (In petals the expression was higher in the distal portion (limb) in comparison to the basal portion (tube). Transgenic plants with reduced PhEXP18 expression levels showed a strong reduction of petal limb size compared to wild-type. Microscopic observations revealed that antisense line petal limb epidermal cells were smaller and showed a reduction in cell wall thickness when compared to wild-type cells. Further analysis revealed that the reduction in thickness of the antisense line cell walls might be attributed to a deficiency in crystalline cellulose.

Proceedings of the XLVII Italian Society of Agricultural Genetics - SIGA Annual Congress
Verona, Italy - 24/27 September, 2003
ISBN 88-900622-4-X

Oral Communication Abstract - S4d

OVEREXPRESSING PHYA GENE CHANGES THE CHERRY PLANT SENSIBILITY TO THE LIGHT SIGNAL

R. MULEO*, C. IACONA**, M.C. INTRIERI*, F. NICESE***, S. MANCUSO***, R. GUERRIERO**, F. LORETI**, B. THOMAS****

*) DiProV, Università della Tuscia (VT), Italy
**) DCDSL, Università di Pisa, Italy
***) Dipartimento di Ortoflorofrutticoltura, Università di Firenze
****) DPGB, HRI, Wellesbourne, Warwick, CV35 9EF, UK

cherry, development, Phytochrome A, Shade Avoidance Response, transgenic plants

Light also acts as a signal of environmental conditions surrounding the plants. There are photoreceptors that function as signal transducers to provide information that controls physiological and morphological responses. Through these pigments, plants have the ability to perceive subtle changes in light composition for initiation of physiological and morphological changes. Known photomorphogenic receptors include phytochrome, cryptochrome and phototropin. Aim of this work was to unravel the effect of phytochrome A overexpression in cherry plants grew under canopy competition, following their reaction to the syndrome of shade avoidance response.

The cherry genotype Colt (P.avium x P.cerasus) was infected with a binary vector systems of Agrobacterium tumefaciens carryng NptII and rice phyA genes, under 35S promoter. From molecular analyses 9 clones resulted positive to the trasformation, and both genes were successfully inserted. All clones expressed the transgenes. The photobiological reactivity of all clones was evaluated in an in vitro screening system. Result showed that the pattern of plantlets development of only four clones was differently regulated by red and far-red light qualities respect to wt.

On the basis of these results, we evaluated the role of the overexpressed phyA gene on to the perception of light qualities in vivo adapted plants: under simulated interaction between plants and light of the 4 clone and wt; while only two clones and wt were studied in real interaction between plants. (Pn), transpiration rate (E) and water use efficiency (WUE) was measured on three fully expanded leaves on every plant of the two different stands under conditions of light saturation (PAR > 1000 µmol m-2s-1).

Plant grew in simulated conditions was surrounding by a light filter, that enriched in red or far-red incoming light direct horizontally on to sapling stem. Mixed stands (that are formed by one clone and wt in chessboard grid) were adopted to grow plants in simulated community canopy, for two years. In both conditions a strong proximity signals were generated. Phenotypic aspects of plant development (stem height, growth, leaf area, branch formation, stomata dimension and formation, chlorophyll content and finally photosynthetic activity) were changed and an intrinsic growth rate was recognised between wt and transformed plants. Net photosynthesis was significantly lower in the transformed PA plants than the wt plants, as the PO plants turned out to be more efficient than the wt ones. The transpiration rate was significantly higher in PO clones compared to wt plants, while water use efficiency (Pn/E) did not differ in all genotypes. Considering the same data after having grouped them in external, middle and internal data according with the position of the plants in the stands, PO plants seemed to be more sensitive to light intensity, as a matter of fact the internal plants showed a higher efficiency in term both of Pn and WUE. A clear relation between the growth and development of transgenic plants and the established concentration of Pfr and the light intensity were also evident, with colt wt and clones plants reacting to neighbours in a divergent dynamic ways. As resulting of these changes, the sensivity to shade and HIR conditions varied among the four clones, ranged from strong shade-tolerant to week shade-tolerant.

This research has been partially supported by: MURST project “Vegetative habit modification of fruit trees through genetic transformation and eco-physiological evaluation of the trangenes”; CNR-Agenzia2000. 

Proceedings of the XLVII Italian Society of Agricultural Genetics - SIGA Annual Congress
Verona, Italy - 24/27 September, 2003
ISBN 88-900622-4-X

Oral Communication Abstract - S4e

ISOLATING TOMATO ROOT MUTANTS TO ASSESS THE FUNCTIONAL BIOLOGY OF ROOT SYSTEMS IN SALINE ENVIRONMENTS

A. MAGGIO, P. DE VITA, P. PALLARA, S. ERRICO

ENEA - National Agency for Innovative Technology, Energy and Environment, C.R. Trisaia, S.S. Jonica Km 419-500, 75026 Rotondella (MT), Italy 

Micro-Tom, mutagenesis, root morphology, root architecture

The role of roots in the short- and long-term adaptation to salt stress is critical. However our understanding of their specific function in saline environments is far from complete. Tomato is an ideal species for the study of plant salt tolerance since it is an economically important crop and well characterized genetic system. Large scale projects to develop a germoplasm resource and a phenotypic catalogue of EMS and fast neutron mutants (http://www.sgn.cornell.edu/mutants/mutants_web/ ) exists. Nevertheless, due to practical problems associated to the inspection of underground organs, the identification of root mutants has been overlooked. To date, only four classes of root mutants have been described in tomato: aerial roots (aer1 and aer2), cottony roots (crt), bushy roots (brt1 and brt2) and dwarf root (drt) mutants. Lack of lateral roots mutants, dgt and gib-1, have been documented, also.  Using the miniature Lycopersicon esculentum cultivar Micro-Tom (Micro tomato) and two different growth systems (polycarbonate panels and hydroponic culture), which both allow visual inspection of the roots, we have started the screening of a large population of mutagenized Micro-Tom for isolating root morphology and architecture mutants.  We are currently screening a transposon tagged population obtained by Dr. Ray Bressan (Purdue University). EMS and fast neutron mutants will be generated for future screening. Preliminary results on the salt tolerance properties of available tomato root mutants will be presented and discussed. 

POSTER

Proceedings of the XLVII Italian Society of Agricultural Genetics - SIGA Annual Congress
Verona, Italy - 24/27 September, 2003
ISBN 88-900622-4-X

Poster Abstract - 4.01

KNOX TRANSCRIPTION FACTORS IN MEDICAGO TRUNCATULA

E. DI GIACOMO, F. SESTILI, G. FRUGIS, D. MARIOTTI

Istituto di Biologia e Biotecnologia Agraria (IBBA), CNR, Area della Ricerca di Roma, Via Salaria Km. 29,300, 00016 Monterotondo Scalo (Roma), IT

transcription factors, Medicago truncatula, KNOX, legume

The model legume Medicago truncatula, a close relative of alfalfa, is a diploid and self-fertile species, with a relatively small genome (4 times that of Arabidopsis thaliana). It is an increasingly popular model plant which is now used worldwide, in particular for studying the molecular genetics of root symbioses.

Homeobox genes are known to play a role in developmental regulation. The knotted-like homeobox (KNOX) genes fall into two classes. The class I KNOX genes like KN1, STM, and KNAT1 are expressed in shoot meristems and loss of function mutations in different species revealed that KNOX genes are involved in the acquisition and maintenance of meristem identity. The function of class II KNOX genes is still undetermined.

In order to characterize KNOX function in Medicago truncatula, constructs for the overexpression of STM and KNAT1 from Arabidopsis and KN1 from maize, under the control of either 35S or a glucocorticoid inducible system (XVE), were obtained. Production of transgenic lines is in progress.

 Through a search in Medicago database, five KNOX genes of M. truncatula have been identified. The corresponding ESTs have been requested to Noble Foundation and the preparation of constructs to overexpress or silence those genes is in progress.

 The expression of endogenous KNOX genes have been investigated by RT-PCR and Northern blotting analysis.

In order to identify possible proteins that interact with KNOX factors in Medicago truncatula, we are going to prepare a cDNA library for two-hybrid analysis. 

Proceedings of the XLVII Italian Society of Agricultural Genetics - SIGA Annual Congress
Verona, Italy - 24/27 September, 2003
ISBN 88-900622-4-X

Poster Abstract - 4.02

The knotted-like class 1 KNAPE1 of peach is triggered and ectopically expressed in leaf affected by the curl disease caused by Taphrina deformans

L. Bruno*, D. Giannino**, A. Chiappetta*, R. Cozza*, A. Tartarini**, D. Mariotti**, A.M. Innocenti*, M.B. Bitonti*

*) Università della Calabria, Dipartimento di Ecologia - Laboratorio di Botanica, Cubo 6B, Arcavacata di Rende, I-87030 Cosenza, Italy 
**) Istituto di Biologia e Biotecnologia Agraria (sez. di Roma), Via Salaria km 29.300, 00016 Monterotondo Scalo (RM)

peach, leaf curl disease, knotted-like genes class 1, cytokinins

In plants, the homeobox containing knotted-like genes (KNOX) class 1 are required for meristem function and determine leaf identity upon down regulation. Most of KNOX class 1 genes are expressed in the shoot apical meristem and excluded from leaf primordia. Mutations of these genes result in a failure to initiate or maintain a SAM, whereas their ectopic expression alters normal leaf morphology, involving a misbalance of hormone metabolism. A knotted-like gene was previously isolated and characterised in peach (KNAPE1) and attributed to class1 on the basis of sequence homology and meristem specific expression pattern. The triggering of KNAPE1 expression was observed in leaves affected by the curl disease, caused by Taphrina deformans, a pathogenic fungus which produces auxin and cytokinin-like compounds. Message localisation was monitored during distinct stages of disease and the ectopic expression was observed in curly sectors, corresponding to areas of histological disorder. This pattern is associated with an indeterminate cell identity, leading to a non coordinate cell proliferation, which may involve a misbalance and/or misallocation of cytokinins. Consequently, immunolocalisation of zeatine was performed and zonation patterns suggest a spatial and temporal relation with the reactivation of KNAPE1. 

Proceedings of the XLVII Italian Society of Agricultural Genetics - SIGA Annual Congress
Verona, Italy - 24/27 September, 2003
ISBN 88-900622-4-X

Poster Abstract - 4.03

FUNCTIONAL ANALYSIS OF THE NF-YB6 GENE FROM ARABIDOPSIS

K. PETRONI, P. GALESI, G. GUSMAROLI, C. TONELLI

Università degli Studi di Milano, Dipartimento di Scienze Biomolecolari e Biotecnologie, Via Celoria 26, 20133 Milano

Transcription factors, CAAT binding proteins, NF-Y, Arabidopsis

In yeast and mammals, the CCAAT box is recognized by NF-Y, a trimer composed of distinct subunits: NF-YA, NF-YB and NF-YC, all required for DNA-binding and encoded by single copy genes (1). In vertebrates, NF-Y promotes and/or stabilises the DNA-binding of many classes of neighbouring transcription factors, sited on adjacent DNA elements, but it has been also proposed a role for NF-Y in chromatin remodelling and nucleosome displacement. Thus, the NF-Y complex could be a key factor in the regulation of the eukaryotic genome during development and differentiation.

A systematic screening of the Arabidopsis thaliana NF-Y homologues led to the identification and cloning of the complete NF-Y family in Arabidopsis, composed of 29 genes: 10 NF-YAs, 10 NF-YBs and 9 NF-YCs, sharing a degree of homology higher than 75% at aminoacidic level with the corresponding yeast and mammal genes (2,3). Their role is largely unknown, except for AtNF-YB9, which corresponds to the Leafy cotyledon1 gene (LEC1), required for embryo maturation and for specification of cotyledon identity in Arabidopsis (4).

Expression profiling of 29 NF-Y transcription factors from Arabidopsis performed by RT-PCR on RNA samples from several organs and developmental stages showed how, inside each family, certain members are expressed ubiquitously, during both vegetative and reproductive growth, while others are transcribed only in specific tissues or developmental stages, in addition to considerable differences in their relative expression levels. In agreement with previous data AtNF-YB9, corresponding to Leafy Cotyledon 1, is induced just in green siliques, during the early embryo developmental stages. 

Based on phylogenetic analysis AtNF-YB6 resulted to be the NF-Y gene more similar to LEC1. Similarly to LEC1, NF-YB6 is expressed in siliques during embryo development and differs from the others at two lysine positions -K78D and K107Q- that are predicted to be important for DNA-binding functions.

Transgenic plants in which the AtNF-YB6 gene is silenced or ectopically expressed have been already produced. T2 overexpression lines show a reduced germination rate and an altered morphology of seedlings. In particular, the hypocotyl and the attachment region of cotyledons are elongated; cotyledons are not fully expanded and often asymmetrical. The first true leaves are characterized by asymmetry both in size and in orientation, while adult plants show distorted rosette leaves. Interestingly, T2 antisense plants show the same phenotype. 

Recently, AtNF-YB6 has been indicated to have a specific role in embryo development, since 7% of RNAi lines for this gene have 30% of defective seeds with embryos blocked at different developmental stages, such as the globular one (5). We are currently analyzing five T-DNA insertion mutants of the AtNFY-B6 gene identified among available collections through database search. One of these carries a T-DNA positioned upstream of ATG, three alleles have a T-DNA insertion inside the coding region, while the fifth one is located downstream the ORF a few base pairs before the polyA signal. 

(1) Maity and de Crombrugghe (1998), Trends Biochem. Sci. 23: 174-178.
(2) Gusmaroli et al., (2001) Gene 264: 173-185.
(3) Gusmaroli et al., (2002) Gene 283:41-48.
(4) Lotan et al. (1998), Cell 93:1195-1205.
(5) Kwong et al. (2003) Plant Cell 15:5-18.

Proceedings of the XLVII Italian Society of Agricultural Genetics - SIGA Annual Congress
Verona, Italy - 24/27 September, 2003
ISBN 88-900622-4-X

Poster Abstract - 4.04

PLANT BODY MODIFICATION IN TWO SUNFLOWER MUTANTS WITH ALTERATIONS IN MERISTEM STRUCTURE

V. MICHELOTTI*, D. BERTINI*, G. CIONINI**, A. CONTI*, M. FAMBRINI*, C. PUGLIESI*

*) Università di Pisa, Dipartimento di Biologia delle Piante Agrarie - Sezione di Genetica, via Matteotti 1/B, 56124 Pisa, Italy
**) Consiglio Nazionale delle Ricerche, Istituto di Biologia e Biotecnologie Agrarie IBBA - Sezione di Pisa, via Moruzzi 1, I-56124 Pisa, Italy

Helianthus annuus, axillary meristems, flowers, phyllotaxy

A vascular plant can be viewed as a branched axis that grows at its tips where two populations of unspecialized cells, the shoot and the root apical meristems, constantly produce organs reiteratively (Clark, 2001). In the post-embryonic vegetative growth, plant form is established by continuos formation of lateral organs and axillary shoots and this phenomenon, is ensured by stem cell homeostasis that requires a negative regulatory feedback loop (Lenhard and Laux, 2003). During the transition of shoot apical meristem from vegetative to reproductive development a change of branching pattern has key importance to differentiate a more complex structure, the inflorescence. Genetic analysis of shoot meristem biology has progressed significantly in recent years especially in the plant model Arabidopsis thaliana (Fletcher, 2002). In this context, the study of mutants affecting the meristem size (with defects in meristem formation and maintenance or characterized by meristem enlargement) is particularly significant (Leyser and Furner, 1992; Barton and Poethig, 1993).

Here we report the morphological and histological analyses of two mutants of sunflower (Helianthus annuus L.) with strong alteration of plant body organization. The missing flowers (mf) mutant show the absence of axillary meristems such as side shoots and flowers (Fambrini et al., 2003), while the stem fasciated (stf) mutant, is characterized by stem fasciation, abnormal phyllotaxy and very short apical internodes. Decapitation experiments and histological analyses indicate that the mf phenotype is the result of a defect in axillary shoot meristem initiation. In addition, we demonstrate that the formation of flowers at the axil of bracts is precluded, in mf/mf plants, by the absence of a subdivion in dyads of floret primordia. On the contrary, a significative enlargement of shoot apical meristem is the primary genetic alteration of the stf mutant.

References
Barton MK., Poethig RS. (1993) Formation of the shoot apical meristem in Arabidopsis thaliana - An analisis of development in the wild type and in the shoot meristemless mutant. Development 119: 823-831.
Clark SE. (2001) Cell signalling at the shoot meristem. Nature Reviews Molecular Cell Biology 2: 276-284.
Fambrini M., Cionini G., Bertini D., Michelotti V., Conti A., Pugliesi C. (2003) MISSING FLOWERS gene controls axillary meristems initiation in sunflower. genesis 36: 25-33.
Fletcher JC. (2002) Shoot and floral meristem maintenance in Arabidopsis. Annual Review Plant Biology 53: 45-66.
Lenhard M., Laux T. (2003) Stem cell homeostasis in the Arabidopsis shoot meristem is regulated by intercellular movement of CLAVATA3 and its sequestration by CLAVATA1. Development 130: 3163-3173.
Leyser HMO., Furner IJ. (1992) Characterization of three shoot apical meristem mutants of Arabidopsis thaliana. Development 116: 397-403.

Proceedings of the XLVII Italian Society of Agricultural Genetics - SIGA Annual Congress
Verona, Italy - 24/27 September, 2003
ISBN 88-900622-4-X

Poster Abstract - 4.05

LEAF SHAPE IN ARABIDOPSIS RELATIVES

P. PIAZZA, M. TSIANTIS

Department of Plant Sciences, University of Oxford, South Parks Road, Oxford OX1 3RB, UK

Arabidopsis, leaf shape, KNOX

All the above-ground organs of a plant originate from a group of cells at the shoot apical meristem (SAM). The SAM serves two main functions: in the central zone, a population, pluripotent stem-like cells is maintained, and in the peripheral zone, lateral organ primordia are initiated.

In Arabidopsis, two KNOX genes, STM and KNAT1 (KNOTTED1-LIKE in ARABIDOPSIS THALIANA1), are expressed in partially overlapping domains of the SAM and are redundantly required for meristem maintenance. Transcripts for both genes, however, are excluded from leaf primordia.

Ectopic expression of STM results in meristem formation on the adaxial surface of the cotyledons and growth arrest [3]. Ectopic expression of KNAT1 not only results in the formation of meristems on leaves but also triggers the development of lobes.

Many closely related species of Arabidopsis thaliana show serrated or lobed leaves thus phenocopying leaves of 35S:KNAT1 plants. This has led to the suggestion that species-specific alterations in KNOX expression patterns result in lobed leaf forms [4].

We aim to test the hypothesis that lobed leaves in Arabidopsis species other than A. thaliana arise as a result of KNOX expression within the leaf and understand the mechanisms specifying persistence of KNOX expression within leaf primordia. Comparative studies between A. thaliana and its dissected leaf relatives should be hugely expedited given the wealth of molecular tools available to study development of A. thaliana.

[1] Long et al., Nature 379, 66–69 (1996)
[2] Byrne et al., Development 129, 1957-1965 (2002)
[3] Williams et al., Bioessays 20, 280–282 (1998)
[4] Bharathan et al., Science 7;296(5574):1858-60 (2002)

Proceedings of the XLVII Italian Society of Agricultural Genetics - SIGA Annual Congress
Verona, Italy - 24/27 September, 2003
ISBN 88-900622-4-X

Poster Abstract - 4.06

DISSECTING GRAPE LEAF DEVELOPMENT AND SENESCENCE AT THE MOLECULAR LEVEL 

C. MOSER*, M. PINDO*, C. SEGALA*, P. FONTANA*, E. BLANZIERI**, P. GATTO*, A. ZAMBONI*, M. BERTAMINI*+, N. NEDUCHENZIAN*+, R. VELASCO* 

*) Istituto Agrario S. Michele a/Adige, S. Michele a/Adige-Trento (+Area Biologia Avanzata; Unità Operativa Viticoltura)
**) Dip. di Informatica e Telecomunicazioni Università di Trento

grape, gene expression, ESTs, arrays, leaf development

Leaf development and senescence are biological processes under tight genetic control. It has been observed that during leaf expansion photosynthetic rates typically increase and vice-versa during leaf senescence they decline. A photosynthetically efficient leaf is important during ripening of grape berries in order to achieve high sugar and secondary metabolites concentrations. High-throughput sequencing of ESTs and gene expression analysis, on a genome-wide scale, have opened new possibilities to shed light on complex biological questions like leaf senescence. 

Following such an approach we have started two years ago the construction and characterization of cDNA libraries from different grape tissues and  the collection of a large number of expressed sequences. The sequence information has been processed via bioinformatics tools in order to estimate redundancy and to assign a putative function to each clone on homology base. Up to now  we sequenced about 8000 ESTs organized in a searchable and soon publicly accessible grapevine database. Clustering of the sequences revealed that about 70 - 85 % of them, are found only in one tissue, suggesting a marked organ specialization, at least for the highly abundant transcripts. A subset of the cDNA clones (ca. 2300 unigenes) have been spotted on high-density nylon filters and probed with RNA isolated from cv.  'Pinot noir' leaves in 5 different developmental stages.  Analysis of gene expression data has been carried out by looking at genes involved in specific metabolic functions and by clustering methods. In addition, transcription profiles have been interpreted in the light of physiological measurements like pigments changes, electron transport activities and total soluble proteins content.

Preliminary results, to be validated and further characterized, identified a group of genes that are up-regulated during leaf senescence.

Proceedings of the XLVII Italian Society of Agricultural Genetics - SIGA Annual Congress
Verona, Italy - 24/27 September, 2003
ISBN 88-900622-4-X

Poster Abstract - 4.07

EPiPHyLLY IN THE VARIANT CLONE EMB-2 OF THE INTERSPECIFIC HYBRID HELIANTHUS ANNUUS X H. TUBEROSUS: EXPRESSION OF A KNOTTED1-LIKE HOMEOBOX GENE AND CYTOKININ DISTRIBUTION IN LEAVES

V. MICHELOTTI*, L. BRUNO**, A. CHIAPPETTA**, M. PETRARULO**, M. FAMBRINI*, M. SALVINI*,****, H. VAN ONCKELEN***, M.B. BITONTI**, C. PUGLIESI*

*) Università di Pisa, Dipartimento di Biologia delle Piante Agrarie - Sezione di Genetica, via Matteotti 1/B, I-56124 Pisa, Italy
**) Università della Calabria, Dipartimento di Ecologia - Laboratorio di Botanica, Cubo 6B, Arcavacata di Rende, I-87030 Cosenza, Italy
***) Department of Biology, University of Antwerp, Belgium
****) Scuola Normale Superiore, Piazza dei Cavalieri, 7 I-56124 Pisa, Italy

homeotic gene, adventitious regeneration, hormone, epiphylly

Structures such as shoots, leaves, embryos and inflorescences may occur upon a leaf or leaf homologue. This phenomenon known as epiphylly is an expression of the morphogenetic potentiality, typical of plant cells (Dickinson, 1978). The flexibility of plant cells has been widely used in vitro tissue cultures which nowadays are considered a support to plant breeding and biotechnology (Jain, 2001). However, the genetic basis of the acquisition of organogenetic and embryogenetic competence are not completely known. It is likely that somatic organogenesis and embryogenesis phenomena in vitro need the same gene regulation hierarchy that control in vivo development of both meristems and zygotic embryos. To verify this hypothesis, an useful plant material are cell lines or mutants that show an high organogenetic and embryogenetic ability without exogenous hormonal requirement (Fambrini et al., 2001).

A somaclonal variant (EMB-2) isolated from in vitro regeneration of the interspecific tetraploid hybrid (2n = 4x = 68) Helianthus annuus x H. tuberosus shows, without hormonal treatments, ectopic formation of epiphyllous shoots and embryos usually arranged in clusters or rows along preexisting veins (Fambrini et al., 2000). This phenotype that is vegetatively propagated resembles some characteristics described in plants with high endogenous cytokinin level or overexpressing Knotted- and/or Leafy Cotyledon-like genes. 

Here we report the expression analysis of a Knotted1-like homeobox gene (Knot1HT) and the pattern of cytokinin accumulation in EMB-2 leaves. The results obtained suggested the involvement of the Knot1HT gene in development of epiphyllic structures. Moreover, we observed a strong signal of trans-zeatin in the parenchimatic cells associated with vascular bundles and along epidermal layer where the cells that resume the mitotic activity are determinated to differentiated the epiphyllic structures. The concomitant localization of cytokinins and Knot1HT transcripts in both the parenchymal cells of vascular bundles and ectopic embryos or shoots, suggest a correlation between these factors and the induction of epiphyllic phenomena.

References
Dickinson TA. (1978) Epiphylly in angiosperms. Botanical Rev. 44: 181-232.
Fambrini M., Cionini G., Bianchi R., Pugliesi C. (2000) Epiphylly in a variant of Helianthus annuus x H. tuberosus induced by in vitro tissue culture. Int. J. Plant Sci. 161: 13-22.
Fambrini M., Fisichella M., Pugliesi C. (2001) Enhanced morphogenetic potential from in vitro regenerated plants of genus Helianthus: an overview. Recent Rec. Develop. Plant Biol.1: 35-54.
Jain SM. (2001) Tissue culture-derived variation in crop improvment. Euphytica 118: 153-166.

Proceedings of the XLVII Italian Society of Agricultural Genetics - SIGA Annual Congress
Verona, Italy - 24/27 September, 2003
ISBN 88-900622-4-X

Poster Abstract - 4.08

GENE EXPRESSION IN BARLEY THROUGH MICROARRAY TECHNOLOGY BY USING NEAR ISOGENIC LINES DERIVED FROM THE “LEAFY LEMMA (LEL)” MUTANT AND THE MALTING QUALITY VARIETY KASKADE

P. Faccioli, F. Rizza, A. Finocchiaro, V. Terzi
 
Istituto Sperimentale per la Cerealicoltura, S.O.P. Fiorenzuola d’Arda, Via San Protaso 302, 29017 Fiorenzuola d’Arda (PC), Italy

barley, morphological mutant, cDNA microarray,  real-time RT-PCR, photosynthetic efficiency

Leafy lemma barley mutant carries recessive alleles of two genes, both of which are necessary to cause the transformation of the lemma into a structure having all characteristics of a vegetative leaf, as shown by SEM analysis. The presence of a sheats, blade, and ligule in the mutant lemma suggests that wild type lemma development is interrupted at a leaf-like stage. Mapping positions of the two lel genes on barley chromosomes 5 and 7 has been determined using F2 populations derived from leafy lemma x Nudinka crosses and an optimized AFLP- based procedure (Castiglioni et al., 1998; Pozzi et al., 2000).

Starting from the cross leafy lemma x Kaskade, near isogenic lines were developed after a six years breeding program: each pair of lines shows the wild type and mutant version of lemma in the same genetic background. The characterization of the isogenic lines was done at three different level. A first characterization of these isogenic lines was done evaluating a set of  agronomic traits in  two years trials. A preliminary evaluation of photosynthetic efficiency of the leafy lemma structure in comparison with other photosynthetic tissues of the plant, like awn and flag leaf was done. Finally, gene expression was studied and compared in leafy lemma, awn and flag leaf from pairs of near isogenic lines using microarray technology (Faccioli et al., 2002). EST clones from a barley leaf cDNA library were arrayed and used to test differences in mRNA expression level in the mutant lemma  in comparison with other photosynthetic tissues of the plant. To support the microarray-based results, the variation in hybridization signals in some clones was confirmed by real time RT-PCR analysis.

References
Castiglioni P., C. Pozzi, M. Heun, K.J. Muller, V.Terzi et al. 1998. An AFLP-based procedure for the efficient mapping of mutations and DNA probes in barley. Genetics 149: 2039-2056.

Pozzi C., P. Faccioli, V. Terzi, A.M. Stanca, S. Cerioli, et al. 2000. Genetics of mutations affecting the development of a barley floral bract. Genetics 154: 1335-1346.

Faccioli P., M.S. Lagonigro, L. De Cecco, A.M. Stanca, R. Alberici, V. Terzi. 2002. Analysis of differential expression of barley ESTs during cold acclimatization using microarray technology. Plant biol. 4: 630-639.

Proceedings of the XLVII Italian Society of Agricultural Genetics - SIGA Annual Congress
Verona, Italy - 24/27 September, 2003
ISBN 88-900622-4-X

Poster Abstract - 4.09

CONSTITUTION OF MAIZE SEGMENTAL BDLs FOR THE FINE MAPPING OF root-ABA1

S. SALVI, M. BELLOTTI, P. LANDI, S. GIULIANI, M.C. SANGUINETI, S. STEFANELLI, S. CONTI, R. TUBEROSA

Department of Agroenviromental Sciences and Technology, University of Bologna, Via Fanin 44, 40137 Bologna, Italy

abscisic acid, maize, positional cloning, QTL, root traits

Root-ABA1 is a quantitative trait locus (QTL) affecting root traits, leaf ABA concentration and grain yield in maize. Root-ABA1 had been previously identified in a mapping population derived from the cross Os420 x IABO78 (Tuberosa et al. 1998, T.A.G. 97:744-755) as a major QTL influencing leaf ABA concentration (L-ABA). The QTL region for high L-ABA (“+” allele) is contributed by Os420 and the QTL region for low L-ABA (“-” allele) by IABO78. Root-ABA1 was mapped on maize chromosome 2 near csu133 on bin 2.04.

Following five backcrosses, backcross derived lines (BDLs) homozygous for the (+/+) or the (-/-) QTL alleles were investigated for L-ABA concentration in a field trial at different water regimes (Landi et al. 2002, M.N.L. 76:7-8). Highly significant differences in L-ABA were detected under both water regimes. Furthermore, the (+/+) and (-/-) hybrids between the BDLs were evaluated for L-ABA, grain yield and other agronomic traits in 2001 and 2002. In all years, highly significant differences were detected for L-ABA. Interestingly, in 2001 the high L-ABA hybrids showed a significantly lower root lodging (44.6%) compared to the low L-ABA hybrids (66.1%). However, the grain yield of the high L-ABA hybrids (41.6 q/ha) was significantly lower than that of the low L-ABA hybrids (54.7 q/ha). Kernel number/plant was the yield component most affected. The results of a pot experiment have shown a significant effect of the QTL on root length, but not on root biomass. Based on these results, we hypothesize that the primary action of the QTL is on root structure which, in turn, may affect directly root lodging and, indirectly via an effect on water status, ABA concentration, ear sterility and yield.

BDLs with contrasting allelic composition at root-ABA1 have also been sampled for RNA extraction at different levels of water stress. Microarray analysis with an oligonucleotide rice array (Affimetrix GeneChipTM) is in progress in collaboration with Syngenta (Giuliani et al., unpublished results).

The positional cloning of root-ABA1 will be attempted starting from a large mapping population produced by crossing Os420 and the Os420 (-/-) BDL carrying the root-ABA1 region (ca. 30-cM long) from IABO78. To improve the resolution around root-ABA1 several SSRs markers (from MaizeDB, http://www.agron.missouri.edu) were tested. Ten SSRs were mapped in the region of interest. In 2002, ca. 1,000 BC6F2 plants were grown. Two SSRs loci (bnlg381 and umc2030) flanking (30-cM) root-ABA1 were used to genotype each plant at the region harbouring root-ABA1. All F2 plants showing recombination events (ca. 200) in the 30-cM interval were selfed and were grown as recombinant BC6F3 families in 2003. After molecular markers analysis, all F3 plants homozygous for a recombination event between the two markers flanking root-ABA1 were selfed. Eventually, all segmental BDLs homozygous for different crossover events at root-ABA1 will be tested for fine mapping and to ascertain to what extent the effects on L-ABA, root traits and yield might be due to one or more linked QTLs.

Based on the results of the fine mapping of segmental BDLs, the microarray analysis, and by exploiting the maize-rice synteny maps we plan to identify a maize physical chromosome region (e.g. BAC contig) carrying the gene/s responsible for root-ABA1. Interestingly, the results of a preliminary survey indicated that the rice region syntenic to maize bin 2.04 harbors root QTLs in several rice populations (Tuberosa et al., unpublished results).

Proceedings of the XLVII Italian Society of Agricultural Genetics - SIGA Annual Congress
Verona, Italy - 24/27 September, 2003
ISBN 88-900622-4-X

Poster Abstract - 4.10

CHARACTERIZATION OF A MAIZE MUTANT AFFECTING EMBRYO AND SEEDLING DEVELOPMENT

M. Colombo*, A. Adamo*, C. Dall’Aglio*, F. Dalla Vecchia**, S. Dolfini***, G. Gavazzi*, A. Giulini*, R. Pilu*, N. Rascio**, G. Consonni*

*) Dipartimento di Produzione Vegetale, Università degli Studi di Milano, Via Celoria 2, 20133 Milan
gabriella.consonni@unimi.it
**) Dipartimento di Biologia, Università degli Studi di Padova, Via Colombo 3, 35121 Padova
***) Dipartimento di Genetica e di Biologia dei Microrganismi, Università degli Studi di Milano, Via Celoria 23, 20133 Milano

maize, developmental mutants, seedling development, cell wall

In maize embryogenesis the coleoptile and first leaves make their appearance during the period from 14 to 20 days after pollination. The coleoptile is formed initially as a ridge of tissue surrounding the central cell stem meristem. The ridge develops more rapidly above then below the central meristem and forms a sheating structure that envelops the stem tip and embryonic leaves. During germination, the plumule and the coleoptile begin to elongate: at first the coleoptile grows faster than the plumule but when it reaches the surface of the soil and is thus exposed to light, it soon ceases to grow and the plumule breaks out from the tip. 

Characterization of the fused leaves (fdl) mutant phenotype suggested that the fdl gene is required during the initial phases of seedling growth. Homozygous fdl seedlings are retarded in their germination when compared to wild-type siblings. Moreover they show regions of fusion between the coleoptile and the first leaf, and between the first and the second leaf. Studies performed at ultrasctructural level have revealed that these fusions involve the cell wall of epidermal cells. The cuticle layer is not present between the two epidermis that appear joined by a single cell wall.

We have analysed the expression of Beta-expansins in the mutant seedlings. Expansins are plant cell wall proteins first discovered in studies of plant cell enlargement as the mediators of acid growth. Expansins are classified in two families, which are called alfa and beta-expansins and many cDNA and genes have been identified in plants. They have unique “loosening” effects on plant cell walls and are thought to have a role in different processes, including cell separation (1). A higher level of ExpB2 mRNA was detected in fdl mutant seedlings during initial phases of seedling growth when leaves are still enrolled inside the coleoptile.
Longitudinal sections of fdl mutant embryos have shown that in the shoot apex coleoptilar cells occupy an ectopic position in the central region, which is normally occupied by leaf primordia. 

Embryo rescue experiments have confirmed that fdl mutation affects the pre-germinative phase of development and have allowed us to assay the effect of exogenous auxins application to cultivated immature embryos. We will present data on the different sensitivity of the mutant embryos to the hormone treatments. 

Taken together these observations suggest that the mutation has a pleiotropic effect on both embryo and seedling growth in maize.  fdl has been located on the long arm of chromosome 7 between UMC 1342 and UMC 1125. Molecular genetic analysis indicate that the mutation is presumably caused by insertion of an Spm element in the fdl locus. Efforts aiming at cloning the gene are underway. 

1. Daniel J. Cosgrove. Enzymes and other agents that enhance cell wall extensibility. Annu. Rev. Plant Physiol. Plant Mol. Biol. 50:391-417 (1999)

Proceedings of the XLVII Italian Society of Agricultural Genetics - SIGA Annual Congress
Verona, Italy - 24/27 September, 2003
ISBN 88-900622-4-X

Poster Abstract - 4.11

Modifications of growth pattern in kiwifruit and cherry induced by the T-DNA genes of Agrobacterium rhizogenes 

R. BIASI*, P. GUTIERREZ*, M. MUGANU*, P. MAGRO**, M. BERNABEI***, E. RUGINI*

*) Dipartimento di Produzione Vegetale, Università degli Studi della Tuscia, Facoltà di Agraria, 01100 Viterbo, Italy
**) Dipartimento di Difesa delle Piante, Università degli Studi della Tuscia, Facoltà di Agraria, 01100 Viterbo, Italy
***) Dipartimento di Tecnologie, Ingegneria dell'Ambiente e delle Foreste, Università degli Studi della Tuscia, Facoltà di Agraria, 01100 Viterbo, Italy

genetic transformation, Agrobacterium rhizogenes, kiwifruit, cherry, vegetative habit, growth pattern, rol genes 

Growth habit of the 5-year-old kiwifruit vines of the transgenic female, cv Hayward, and the male selection GTH, and the cherry rootstock Colt, has been evaluated in an authorised open field since 1998. 

Kiwifruit (Actinidia deliciosa A. Chev.), genetically modified, were obtained in vitro from leaf explants co-cultivated with A. tumefaciens carrying the rolABC and rolB genes of A. rhizogenes under natural promoter (Rugini et al, 1991). The introduction of the rolB does not seems to modify plant morphology, while the simultaneous introduction of the three associated genes, rolABC, induced plant modification regarding the phenology stages and plant architecture in both female and male vines. Bud burst was delayed, resulting in a lower number of lateral shoots compared to untransformed control plants; however, the basal buds always developed normally, showing a marked basitony behaviour. The lateral shoots in transgenic plants, beside having shortened internodes, carried leaves in upright position, with a reduced lamina surface and increased thickness. In addition, the plants showed an increased of water retention during a waterless stress period, which could be due to a modified hydraulic conductivity in the different xylem anatomy, since the elements were smaller and pits were extended in all cell walls of vessels.  Nitrogen content in the leaves was also increased in almost all somaclones tested.

Cherry rootstock "Colt" (Prunus avium x Prunus pseudocerasus) plants were also produced in vitro from transgenic roots induced by A. rhizogenes wild type (Gutierez-Pesce et al., 1998). The plants showed variability in architecture of the growth pattern within the regenerated somaclones: 50% did not show the characteristic of "hairy root phenotype" so their growth pattern was similar to untransformed control plants; the other 50% of the transgenic somaclones showed a reduced total growth (height and diameter) resulting in a spur habit respect to control plants, i.e. marked short internodes and reduced vigour, a leaf area drastically reduced, as well as length of the petiole and fresh and dry weight reduction, while chlorophyll and nitrogen content did not change in any of the transgenic somaclones. Regarding flowering, all replicates of one transgenic somaclone (somaclone A) never produced flowers.  The period of blossom in the rest of the transgenic somaclones was delayed by several days, while the morphology or the flowers was similar to the untransformed control plants. Delayed was also the rest period. When transgenic rootstocks were grafted with normal scion, in some case, the bimember plant size resulted reduced. The rooting ability of both herbaceous and hardwood cutting of transgenic somaclones with "hairy root phenotype" aspect, was unexpectedly drastically low.

Proceedings of the XLVII Italian Society of Agricultural Genetics - SIGA Annual Congress
Verona, Italy - 24/27 September, 2003
ISBN 88-900622-4-X

Poster Abstract - 4.12

SOMATIC HYBRIDIZATION BETWEEN EGGPLANT (SOLANUM MELONGENA) AND WILD ARBOREOUS SOLANUM SPECIES

F. PISANI, D. NIGRIS, L. BORGATO, A. FURINI

Dip. Scientifico e Tecnologico, Università degli Studi di Verona, Strada Le Grazie 15, 37134 Verona

Solanum is one of the largest genera of vascular plants consisting of more than 1500 described species. Therefore, it represents a source of variability that may be explored for the genetic improvement of many different traits. The selection of somatic hybrids between the herbaceous cultivated eggplant and wild Solanum species with arboreous growth habit is the aim of this study. S. linnaeanum, S. marginatum, S. virginianum and S. campylacanthum have been selected for their woody growth. Although a phylogenetic study by AFLP molecular markers has demonstrated genetic affinity of these species with the cultivated S. melongena, they showed sexual incongruity. 

For the heterokaryon selection, S. melongena was genetically transformed to introduce the neomycinphosphotransferase II and the bar genes as selectable markers, while the other partners were biotinylated. After the fusion process achieved by electrofusion, protoplasts were labelled with magnetic microbeads coupled to antibiotin, in this way hybrid cells were resistant to kanamycin and phosphinothricin (PPT) and magnetically labelled. They were selected by passing the fusion mixture into the MACS (magnetic cell sorter) and then incubating the magnetic fraction on kanamycin- or PPT-containing medium to eliminate the homokaryotic cells derived from the kanamycin and PPT susceptible biotin-labelled parent. Regeneration from microcalli may result not only in plants characterized by hybrid vigor but also by different plant architecture with arborescent or shrubby habitus and the new varieties may have perennial instead of annual fruit production.

Proceedings of the XLVII Italian Society of Agricultural Genetics - SIGA Annual Congress
Verona, Italy - 24/27 September, 2003
ISBN 88-900622-4-X

Poster Abstract - 4.13

GENETIC DISSECTION OF SHOOT DEVELOPMENT IN BARLEY (HORDEUM VULGARE L.): AN EST-BASED CANDIDATE GENE STRATEGY

L. NICOLOSO*, C. POZZI**, F. SALAMINI*,***, L. ROSSINI*

*) University of Milan, Dipartimento di Produzione Vegetale, V. Celoria 2, 20133 Milan, Italy 
**) Fondazione Parco Tecnologico Padano, V. Haussmann 11, 26900 Lodi, Italy
***) Max Planck Institute for Plant Breeding Reasearch, Carl-von-Linné Weg 10, 50829 Koeln, Germany

Hordeum vulgare, developmental mutants, candidate gene approach, linkage maps, SNP

A variety of barley mutants affecting shoot and floral architecture have been collected at the Max Planck Institute in Koeln. At present 35 developmental mutants have been positioned on the established high density molecular linkage map of barley (1, 2). Isolation of the corresponding genes will provide insight into the molecular mechanisms underlying development of the Triticeae. In barley, positional cloning is hampered by the low degree of polymorphism and the large size of the genome (4.873 Mbp). However, map-based cloning can be accelerated by the identification of candidate genes (CGs), as demonstrated by the association of the barley Hooded phenotype with a mutation in the homeobox gene Bkn3 (3).

In order to identify barley CGs for the mutants under study in our group, we initially compiled a list of 157 regulatory genes known for their key-role in shoot development of Arabidopsis, maize, rice etc. These sequences were used as queries in similarity searches on the barley EST database (http://www.ncbi.nlm.nih.gov/), uncovering a total number of 423 homologous ESTs, putatively assigned to different transcription factor families.

In the next step of this strategy, candidate ESTs are mapped and putative cosegregation with developmental mutants is analysed. SNPs are the ideal tool for EST mapping and can be identified in silico (isSNPs) by comparison of ESTs corresponding to the same gene but derived from different genotypes. This approach lead to the identification of 144 barley isSNPs in our candidate ESTs. Prior to mapping, these isSNPs are experimentally validated on the parents of available  mapping populations. If polymorphism for a given gene is not detected on these lines, the screening proceeds on the 67 wild barley lines already utilised for mapping in our group (1).

Castiglioni et al (1998) Genetics 149: 2039
Pozzi et al. (2003) Heredity 90: 39
Müller et al. (1995) Nature 374: 727

Proceedings of the XLVII Italian Society of Agricultural Genetics - SIGA Annual Congress
Verona, Italy - 24/27 September, 2003
ISBN 88-900622-4-X

Poster Abstract - 4.14

A RCC1-LIKE GENE CONTROLS GROWTH AND DEVELOPMENT IN ARABIDOPSIS

V. VALIANTE*, F. CONDIGLIO*, S. GRILLO*, R. SERRANO**, A. LEONE***

*) Institute of Plant Genetics (CNR-IGV) - Research dDvision of Portici, Naples, Italy
**) Instituto de Biologia  Molecular y Celular de Plantas, CSIC, Valencia, Spain
***) Dept. of Pharmaceutical Sciences, University of Salerno, Italy

Arabidopsis, plant growth and development, cell cycle gene regulation 

Beta-propellers are emerging as novel protein domains promoting protein-protein interaction and binding to ligands, sugars, prosthetic groups and ions -propellers modules are formed by a variety of divergent consensus aminoacid sequences, found in polypeptides involved in different cellular activities (e.g. WD motifs of G protein  subunit, kelch repeat proteins etc. 

An Arabidopsis gene encoding a putative -propeller protein was identified as able to restore growth of the osmotic unstable mpk-ppz1- yeast mutant in absence of sorbitol. Besides the presence of seven RCC1-like motifs, the protein contains also a conserved putative cyclin binding domain.

A functional gain and loss approach was used to get insights into the role of this plant gene. Compared to wild type plants, T4 over-expressing plants had a stunted vegetative growth and a reduced number of flowers and siliques. Antisense plants were very vigorous, with a larger rosette and leaf size and a significant higher number of inflorescence and siliques. These data suggest that the gene may act as a negative regulator of Arabidopsis plant growth, as further confirmed by phenotypic analysis of a mutant from the Salk knock-out collection. Cyclin, ICK and CDK genes were differently regulated in sense and antisense plants, indicating that the gene might be involved in plant cell cycle.