HISTONE DEACETYLASE19 Controls Ovule Number Determination and Transmitting Tract Differentiation.

The gynoecium is critical for the reproduction of flowering plants as it contains the ovules and the tissues that foster pollen germination, growth, and guidance. These tissues, known as the reproductive tract (ReT), comprise the stigma, style, and transmitting tract (TT). The ReT and ovules originate from the carpel margin meristem (CMM) within the pistil. SHOOT MERISTEMLESS (STM) is a key transcription factor for meristem formation and maintenance. In all above-ground meristems, including the CMM, local STM downregulation is required for organ formation. However, how this downregulation is achieved in the CMM is unknown. Here, we have studied the role of HISTONE DEACETYLASE 19 (HDA19) in Arabidopsis (Arabidopsis thaliana) during ovule and ReT differentiation based on the observation that the hda19-3 mutant displays a reduced ovule number and fails to differentiate the TT properly. Fluorescence-activated cell sorting coupled with RNA-sequencing revealed that in the CMM of hda19-3 mutants, genes promoting organ development are downregulated while meristematic markers, including STM, are upregulated. HDA19 was essential to downregulate STM in the CMM, thereby allowing ovule formation and TT differentiation. STM is ectopically expressed in hda19-3 at intermediate stages of pistil development, and its downregulation by RNA interference alleviated the hda19-3 phenotype. Chromatin immunoprecipitation assays indicated that STM is a direct target of HDA19 during pistil development and that the transcription factor SEEDSTICK is also required to regulate STM via histone acetylation. Thus, we identified factors required for the downregulation of STM in the CMM, which is necessary for organogenesis and tissue differentiation.

Cell wall modifications by α-XYLOSIDASE1 are required for control of seed and fruit size in Arabidopsis.

Cell wall modifications are of pivotal importance during plant development. Among cell wall components, xyloglucans are the major hemicellulose polysaccharide in primary cell walls of dicots and non-graminaceous monocots. They can connect the cellulose microfibril surface to affect cell wall mechanical properties. Changes in xyloglucan structure are known to play an important role in regulating cell growth. Therefore, the degradation of xyloglucan is an important modification that alters the cell wall. The α-XYLOSIDASE1 (XYL1) gene encodes the only α-xylosidase acting on xyloglucans in Arabidopsis thaliana. Here, we showed that mutation of XYL1 strongly influences seed size, seed germination, and fruit elongation. We found that the expression of XYL1 is directly regulated in developing seeds and fruit by the MADS-box transcription factor SEEDSTICK. We demonstrated that XYL1 complements the stk smaller seed phenotype. Finally, by atomic force microscopy, we investigated the role of XYL1 activity in maintaining cell stiffness and growth, confirming the importance of cell wall modulation in shaping organs.

Gynoecium size and ovule number are interconnected traits that impact seed yield.

Angiosperms form the largest group of land plants and display an astonishing diversity of floral structures. The development of flowers greatly contributed to the evolutionary success of the angiosperms as they guarantee efficient reproduction with the help of either biotic or abiotic vectors. The female reproductive part of the flower is the gynoecium (also called pistil). Ovules arise from meristematic tissue within the gynoecium. Upon fertilization, these ovules develop into seeds while the gynoecium turns into a fruit. Gene regulatory networks involving transcription factors and hormonal communication regulate ovule primordium initiation, spacing on the placenta, and development. Ovule number and gynoecium size are usually correlated and several genetic factors that impact these traits have been identified. Understanding and fine-tuning the gene regulatory networks influencing ovule number and pistil length open up strategies for crop yield improvement, which is pivotal in light of a rapidly growing world population. In this review, we present an overview of the current knowledge of the genes and hormones involved in determining ovule number and gynoecium size. We propose a model for the gene regulatory network that guides the developmental processes that determine seed yield.

Cytokinin response factors integrate auxin and cytokinin pathways for female reproductive organ development.

The developmental programme of the pistil is under the control of both auxin and cytokinin. Crosstalk between these factors converges on regulation of the auxin carrier PIN-FORMED 1 (PIN1). Here, we show that in the triple transcription factor mutant cytokinin response factor 2 (crf2) crf3 crf6 both pistil length and ovule number were reduced. PIN1 expression was also lower in the triple mutant and the phenotypes could not be rescued by exogenous cytokinin application. pin1 complementation studies using genomic PIN1 constructs showed that the pistil phenotypes were only rescued when the PCRE1 domain, to which CRFs bind, was present. Without this domain, pin mutants resemble the crf2 crf3 crf6 triple mutant, indicating the pivotal role of CRFs in auxin-cytokinin crosstalk.

Alternative Splicing Generates a MONOPTEROS Isoform Required for Ovule Development.

The plant hormone auxin is a fundamental regulator of organ patterning and development that regulates gene expression via the canonical AUXIN RESPONSE FACTOR (ARF) and AUXIN/INDOLE-3-ACETIC ACID (Aux/IAA) combinatorial system. ARF and Aux/IAA factors interact, but at high auxin concentrations, the Aux/IAA transcriptional repressor is degraded, allowing ARF-containing complexes to activate gene expression. ARF5/MONOPTEROS (MP) is an important integrator of auxin signaling in Arabidopsis development and activates gene transcription in cells with elevated auxin levels. Here, we show that in ovules, MP is expressed in cells with low levels of auxin and can activate the expression of direct target genes. We identified and characterized a splice variant of MP that encodes a biologically functional isoform that lacks the Aux/IAA interaction domain. This MP11ir isoform was able to complement inflorescence, floral, and ovule developmental defects in mp mutants, suggesting that it was fully functional. Our findings describe a novel scenario in which ARF post-transcriptional regulation controls the formation of an isoform that can function as a transcriptional activator in regions of subthreshold auxin concentration.

MADS-Box and bHLH Transcription Factors Coordinate Transmitting Tract Development in Arabidopsis thaliana.

The MADS-domain transcription factor SEEDSTICK (STK) controls several aspects of plant reproduction. STK is co-expressed with CESTA (CES), a basic Helix-Loop-Helix (bHLH) transcription factor-encoding gene. CES was reported to control redundantly with the brassinosteroid positive signaling factors BRASSINOSTEROID ENHANCED EXPRESSION1 (BEE1) and BEE3 the development of the transmitting tract. Combining the stk ces-4 mutants led to a reduction in ovule fertilization due to a defect in carpel fusion which, caused the formation of holes at the center of the septum where the transmitting tract differentiates. Combining the stk mutant with the bee1 bee3 ces-4 triple mutant showed an increased number of unfertilized ovules and septum defects. The transcriptome profile of this quadruple mutant revealed a small subset of differentially expressed genes which are mainly involved in cell death, extracellular matrix and cell wall development. Our data evidence a regulatory gene network controlling transmitting tract development regulated directly or indirectly by a STK-CES containing complex and reveal new insights in the regulation of transmitting tract development by bHLH and MADS-domain transcription factors.

REM34 and REM35 Control Female and Male Gametophyte Development in Arabidopsis thaliana.

The REproductive Meristem (REM) gene family encodes for transcription factors belonging to the B3 DNA binding domain superfamily. In Arabidopsis thaliana, the REM gene family is composed of 45 members, preferentially expressed during flower, ovule, and seed developments. Only a few members of this family have been functionally characterized: VERNALIZATION1 (VRN1) and, most recently, TARGET OF FLC AND SVP1 (TFS1) regulate flowering time and VERDANDI (VDD), together with VALKYRIE (VAL) that control the death of the receptive synergid cell in the female gametophyte. We investigated the role of REM34, REM35, and REM36, three closely related and linked genes similarly expressed in both female and male gametophytes. Simultaneous silencing by RNA interference (RNAi) caused about 50% of the ovules to remain unfertilized. Careful evaluation of both ovule and pollen developments showed that this partial sterility of the transgenic RNAi lines was due to a postmeiotic block in both female and male gametophytes. Furthermore, protein interaction assays revealed that REM34 and REM35 interact, which suggests that they work together during the first stages of gametogenesis.