Research

The developmental potentials of plant meristems

In plants, organogenesis is maintained throughout life by specialized tissues called meristems (from merizein – Greek, to divide). There are several analogies between meristems and animal stem cells, in particular, the asymmetric divisions that maintain meristematic cells on one hand and initiate plant organs on the other. Apical plan meristems are relatively large, made of nearly 100 cells and maintain relatively uniform shape and dimensions even when different organs are produced.

While maintenance mechanisms of apical meristems by feedback loops governing the WUSCHEL homeobox expression are fairly understood, it is not clear how meristem cells are changing upon developmental switches. Such developmental switches may alter the organs produced by the apical meristem, e.g. leaves or bracts, the branching shoots, e.g. side shoot or a stolon, or even the meristem proper that can be maintained or turn into a flower. We use the tomato shoot to understand the genetic and molecular mechanisms that underlie these switches in general, and to follow several hormones, auxin, cytokinin and florigen in particular. Significantly, these are general growth hormones and their role in specific developmental switches is not understood. Several ongoing studies addressing different aspects of apical meristems biology are outlined.

Formation of lateral organs at the flanks of the meristem

To uncover the molecular and physiological mechanisms that balance differentiation and indeterminate growth, we use the leafless (lfs) tomato mutant. lfs, a transcription factor mutant exhibits precocious developmental arrest and a failure to form leaves. By comparing the expression profiles of lfs and the wild-type tomato apices, we observed up-regulation of SLR and PAP, two AUX/IAA genes that negatively regulate auxin signaling. Expression of non-degradable SLR leads to recapitulation of the lfs phenotype. These findings suggest that LFS is involved in auxin signaling and may mediate the meristem’s ability to differentiate lateral organs in response to the hormone.

A. leafless plants (right) are small, under-developed, lack cotyledons and usually lack leaves (cf. wild type plant at the same age on the left).

B. Expression of SLR in WT and in lfs.

C. Plant expressing a mutated version of SLR under the LFS promoter recapitulates the lfs phenotype.

Regulation of tomato floral transition by LTM, a KELCH protein lost in crucifers

During vegetative development, the shoot apical meristem (SAM) maintains a relatively constant shape and size. Transition from a vegetative to reproductive phase is first evident by enlargement and doming of the SAM, however the mechanisms that drive this dramatic physical change of the SAM are unknown. In a large scale screen for mutants with delayed flowering we identified a new tomato gene, late termination (ltm) that regulates floral transition in both primary and sympodial meristems. A double mutant analysis of ltm and the tomato Florigen gene single flower truss (sft) resulted in an additive late flowering phenotype, suggesting that LTM is not part of the SFT flowering pathway. Surprisingly, during the vegetative phase ltm SAM has a domed shape, unlike the flat SAM of vegetative WT. Transcriptomes of dissected ltm and WT meristems in different vegetative stages uncovered a surprising trend; most of the genes that were up regulated in ltm apices were genes that under normal conditions are activated towards floral transition. Significantly, none of these genes are precociously up regulated in mutants with large meristems such as clavata. Of these genes, SELF PRUNING (SP) is a CETS protein that is up regulated toward floral transition where it functions antagonistically to the flowering process. In agreement, RNA In Situ hybridization showed that SP is expressed at the vegetative SAM of ltm but not in WT.

Ectopic expression of SP resulted in delayed floral transition as well as induction of a domed vegetative meristem. ltm sp plants flowered earlier than ltm, however later than WT plants. Taken together, upregulation of SP is only partially responsible for the delayed flowering of ltm suggesting that other genes miss regulated in ltm may function as floral transition regulators. LTM encodes for a nuclear localized protein containing five Kelch domains. It is strongly expressed at the border between the apex and emerged leaves and has a weak expression in the vegetative SAM that expands during floral transition. Using a BLAST survey, several close homologs of the protein were identified in other plant species however not in Brassicaceae plants. Similarly, three of the genes that were strongly up regulated in ltm are also missing from these genomes. Thus, LTM is part of a floral regulation pathway evolutionally lost from the Arabidopsis genome; whether it is required to balance between SAM termination by floral signals and the need to maintain growth in flowering perennial plants awaits further analyses.

Primery vs. side shoots

The primary shoot meristem (PSM) is gradually formed from the apical part of the developing zygote. Axillary meristems initiate at the boundary of leaf primordia and the stem to form side shoots (SS). Both types of meristems will generate stem, leaves and flowers. Each meristem on the same plant will differentiate into flowers at different times, even though the flowering hormone florigen is systemic and emitted from mature leaves. To uncover the molecular mechanisms underlying the differences between the two meristem types an RNA Expression Dataset of primary and side shoots was constructed from RNA of Primary Shoot Meristem (PSM) and 3 types of Side Shoot meristems (SS); leaf axil -1, -3 and +3 (A)

In broad terms, side shoots have high expression of members that belong to several groups of transcription factors (TF) such as TCP and SPL that may repress SS growth. In addition, several TFs from MADS box family, that include multiple homeotic regulators, show quantitative differences between the two meristem classes (B). Likewise, clear differences in expression of genes directing Cytokinin metabolism (C) were evident. Could these represent novel components that differentiate meristem fate?

The broad effects of the Florigen complex

The current model for long distance regulation of flowering relies on the formation of the Florigen in sensing organs (leaves) and its movement towards its site of response- the SAM. Recent studies in tomato, Arabidopsis and rice provided compelling evidence that the product(s) of the FLOWERING LOCUS T (FT) or its tomato homolog, the SFT gene, is florigen. In the SAM, the florigen co-operate with its "receptor", a b-ZIP transcription factor to promote the transition from vegetative to reproductive state. The b-ZIP gene of tomato, the SPGB2, has two spliced forms. When each of these forms is expressed together with SFT the plants are stunted, although this synergistic interaction is more pronounced when the SPGB2 form maintains its first intron (iSPGB2) (A). Significantly, this interaction also occurs when a scion expressing Florigen is grafted with rootstock expressing a b-ZIP receptor, the FDP protein (B).

(A) The interactions between the two forms of SPGB2 and SFT. The numbers of leaves till the first inflorescence are marked. The arrows pointing upon the first inflorescence. (B) An 35S:SFT scion grafted on sft 35:FDP rootstock. Note the big leaves of the donor and the minute leaves of the recipient rootstock. Displayed are 35:FDP recipient rootstocks where the donor was maintained (left) or removed three weeks before the picture was taken (right). A close up of a shoots from the 35:FDP rootstocks is exhibit.

Hidden evolutionary potentials: making tubers in tomato

Plant storage organs constitute a vegetative alternative to sexual reproduction. Storage organs vary in their origins, as manifested by the swelling roots of carrots and sweet potato versus the expanded hypocotyls of beets versus the familiar potato tubers. Potato tubers develop from the swelling tips of specialized underground lateral shoots called stolons. In contrast, axillary meristems of the closely related tomato never produce tubers but only aerial shoots. Intriguingly, the ectopic expression of LONELY GUY1 (LOG1) or IPT, genes encoding cytokinin (CK) biosinthetic enzymes, triggers the formation of small, sessile and aerial tomato tubers at the axiles of cotyledons, leaves or leaflets (A, B). The tomato tubers are identical to aerial, sessile, mini-tubers produced by potato plants grown under a variety of physiological conditions and reveal a hidden evolutionary potential unleashed by cytokinin signaling and suggest that other meristems also carry evolutionarily suppressed potentials (C).