SHOOT MERISTEMLESS Research Articles

Mechanical stress contributes to the expression of the STM homeobox gene in Arabidopsis shoot meristems.

The role of mechanical signals in cell identity determination remains poorly explored in tissues. Furthermore, because mechanical stress is widespread, mechanical signals are difficult to uncouple from biochemical-based transduction pathways. Here we focus on the homeobox gene SHOOT MERISTEMLESS (STM), a master regulator and marker of meristematic identity in Arabidopsis. We found that STM expression is quantitatively correlated to curvature in the saddle-shaped boundary domain of the shoot apical meristem. As tissue folding reflects the presence of mechanical stress, we test and demonstrate that STM expression is induced after micromechanical perturbations. We also show that STM expression in the boundary domain is required for organ separation. While STM expression correlates with auxin depletion in this domain, auxin distribution and STM expression can also be uncoupled. STM expression and boundary identity are thus strengthened through a synergy between auxin depletion and an auxin-independent mechanotransduction pathway at the shoot apical meristem.

Alternate wiring of a KNOXI genetic network underlies differences in leaf development of A. thaliana and C. hirsuta

Two interrelated problems in biology are understanding the regulatory logic and predictability of morphological evolution. Here, we studied these problems by comparing Arabidopsis thaliana, which has simple leaves, and its relative, Cardamine hirsuta, which has dissected leaves comprising leaflets. By transferring genes between the two species, we provide evidence for an inverse relationship between the pleiotropy of SHOOTMERISTEMLESS (STM) and BREVIPEDICELLUS (BP) homeobox genes and their ability to modify leaf form. We further show that cis-regulatory divergence of BP results in two alternative configurations of the genetic networks controlling leaf development. In C. hirsuta, ChBP is repressed by the microRNA164A (MIR164A)/ChCUP-SHAPED COTYLEDON (ChCUC) module and ChASYMMETRIC LEAVES1 (ChAS1), thus creating cross-talk between MIR164A/CUC and AS1 that does not occur in A. thaliana. These different genetic architectures lead to divergent interactions of network components and growth regulation in each species. We suggest that certain regulatory genes with low pleiotropy are predisposed to readily integrate into or disengage from conserved genetic networks influencing organ geometry, thus rapidly altering their properties and contributing to morphological divergence.

HANABA TARANUregulates the shoot apical meristem and leaf development in cucumber (Cucumis sativusL.)

The shoot apical meristem (SAM) is essential for continuous organogenesis in higher plants, while the leaf is the primary source organ and the leaf shape directly affects the efficiency of photosynthesis. HANABA TARANU (HAN) encodes a GATA3-type transcription factor that functions in floral organ development, SAM organization, and embryo development in Arabidopsis, but is involved in suppressing bract outgrowth and promoting branching in grass species. Here the function of the HAN homologue CsHAN1 was characterized in cucumber, an important vegetable with great agricultural and economic value. CsHAN1 is predominantly expressed at the junction of the SAM and the stem, and can partially rescue the han-2 floral organ phenotype in Arabidopsis. Overexpression and RNAi of CsHAN1 transgenic cucumber resulted in retarded growth early after embryogenesis and produced highly lobed leaves. Further, it was found that CsHAN1 may regulate SAM development through regulating the WUSCHEL (WUS) and SHOOT MERISTEMLESS (STM) pathways, and mediate leaf development through a complicated gene regulatory network in cucumber.

Overexpression of ARBORKNOX1 delays the differentiation of induced somatic sector analysis (ISSA) derived xylem fibre cells in poplar stems

ARBORKNOX1 (ARK1), a poplar functional homolog of the Arabidopsis Class-I KNOTTED- like homeobox (KNOXI) transcription factor SHOOT MERISTEMLESS (STM), has been shown to have a role in maintaining meristematic cells in an undifferentiated state. Studies in which this gene was ectopically overexpressed identified a delay in the differentiation of cambial derivatives. Overexpression of ARK1 in poplar stem sectors, using the induced somatic sector analysis (ISSA) experimental system, was undertaken to further investigate its role in the differentiation of xylem fibre cells. ISSA involves the in vivo transformation of a small number of cambial cells in the stem with a gene of interest giving rise to a discreet transgenic tissue sector that can be directly compared to adjacent non-transgenic fibres to elucidate gene function. Phenotypes obtained were comparable with the proposed role of ARK1, where a delay in the development of the cell wall in differentiating fibre cells of active cambia was observed. Additional evidence suggests a role for ARK1 in the positive regulation of cell viability in the cambium. The significance of dormancy signals in the context of differentiation was also highlighted, as some transgenic fibres failed to fully differentiate prior to the onset of dormancy. Our findings are discussed in the context of wood formation and the use of ISSA as an experimental system.

Overcoming recalcitrance - Auxin response factor functions in plant regeneration

De novo meristem formation in tissue culture critically depends on the correct organization of hormonal domains, which is followed by expression shoot meristem pattern genes. The genetic basis of plant regeneration is fragmentary, but mutant studies demonstrate that signaling through MONOPTEROS (MP)/AUXIN RESPONSE FACTOR 5 is necessary for the formation of shoots from Arabidopsis calli. Most strikingly, variants of MP, uncoupling MP activity from negative regulation by Aux/IAA proteins, showed that MP is also sufficient for promoting de novo shoot formation even from normally recalcitrant tissues. In this function MP acts through pathways involving the homeobox transcription factor SHOOT MERISTEMLESS (STM) and AP2 domain transcription factor CYTOKININ RESPONSE FACTOR2 (CRF2). Our findings provide an entry point to better address the molecular genetics underlying divergent regeneration properties and demonstrate the potential of ARF-derived constructs as novel genetic tools to develop high frequency regeneration systems in recalcitrant explants and species

Developmental morphology of flattened shoots in Dalzellia ubonensis and Indodalzellia gracilis with implications for the evolution of diversified shoot morphologies in the subfamily Tristichoideae (Podostemaceae).

Podostemaceae is a unique family of aquatic angiosperms that grow in swift-running water on rock surfaces in the tropics. Their plant bodies show a remarkable adaptation: the main plant body is mostly creeping or flattened, or in extreme cases foliose, functioning as an adhering and photosynthetic organ. In the subfamily Podostemoideae, the root is foliose, whereas in the subfamily Tristichoideae, the shoot is foliose. An evolutionary scenario for the foliose root has already been proposed, but that for the foliose shoot remains to be addressed. Shoots of Indodalzellia gracilis and Dalzellia ubonensis (subfamily Tristichoideae) were observed using light microscopy and scanning electron microscopy. Gene expression patterns of orthologs of marker genes for the shoot apical meristem, i.e., SHOOT MERISTEMLESS and WUSCHEL, in D. ubonensis were analyzed. When very young, the phyllotaxis is tristichous in both genera: a set of one dorsal and two marginal leaves forms. When the shoot branches, extra-axillary buds of two subsequent marginal leaves form as new (lateral) shoots, and the original shoot stops growing; this growth pattern is called sympodial branching. Due to zonal growth in the common zone just below the original and lateral shoot apices, flattened or foliose shoots result. The expression patterns of DuSTM and DuWUS in the shoot apices of Dalzellia were similar to those published for Terniopsis. The foliose shoots of Indodalzellia and Dalzellia evolved as a result of congenital fusion among several original and lateral branches, each of which grows separately in other Tristichoideae.

Transcriptional, posttranscriptional, and posttranslational regulation of SHOOT MERISTEMLESS gene expression in Arabidopsis determines gene function in the shoot apex.

The activity of SHOOT MERISTEMLESS (STM) is required for the functioning of the shoot apical meristem (SAM). STM is expressed in the SAM but is down-regulated at the site of leaf initiation. STM is also required for the formation of compound leaves. However, how the activity of STM is regulated at the transcriptional, posttranscriptional, and posttranslational levels is poorly understood. We previously found two conserved noncoding sequences in the promoters of STM-like genes across angiosperms, the K-box and the RB-box. Here, we characterize the function of the RB-box in Arabidopsis (Arabidopsis thaliana). The RB-box, along with the K-box, regulates the expression of STM in leaf sinuses, which are areas on the leaf blade with meristematic potential. The RB-box also contributes to restrict STM expression to the SAM. We identified FAR1-RELATED SEQUENCES-RELATED FACTOR1 (FRF1) as a binding factor to the RB-box region. FRF1 is an uncharacterized member of a subfamily of four truncated proteins related to the FAR1-RELATED SEQUENCES factors. Internal deletion analysis of the STM promoter identified a region required to repress the expression of STM in hypocotyls. Expression of STM in leaf primordia under the control of the JAGGED promoter produced plants with partially undifferentiated leaves. We further found that the ELK domain has a role in the posttranslational regulation of STM by affecting the nuclear localization of STM.

Class I KNOX transcription factors promote differentiation of cambial derivatives into xylem fibers in the Arabidopsis hypocotyl.

The class I KNOX transcription factors SHOOT MERISTEMLESS (STM) and KNAT1 are important regulators of meristem maintenance in shoot apices, with a dual role of promoting cell proliferation and inhibiting differentiation. We examined whether they control stem cell maintenance in the cambium of Arabidopsis hypocotyls, a wood-forming lateral meristem, in a similar fashion as in the shoot apical meristem. Weak loss-of-function alleles of KNAT1 and STM led to reduced formation of xylem fibers - highly differentiated cambial derivatives - whereas cell proliferation in the cambium was only mildly affected. In a knat1;stm double mutant, xylem fiber differentiation was completely abolished, but residual cambial activity was maintained. Expression of early and late markers of xylary cell differentiation was globally reduced in the knat1;stm double mutant. KNAT1 and STM were found to act through transcriptional repression of the meristem boundary genes BLADE-ON-PETIOLE 1 (BOP1) and BOP2 on xylem fiber differentiation. Together, these data indicate that, in the cambium, KNAT1 and STM, contrary to their function in the shoot apical meristem, promote cell differentiation through repression of BOP genes.

Irrepressible MONOPTEROS/ARF5 promotes de novo shoot formation.

In vitro regeneration of complete organisms from diverse cell types is a spectacular property of plant cells. Despite the great importance of plant regeneration for plant breeding and biotechnology, its molecular basis is still largely unclear and many important crop plants have remained recalcitrant to regeneration. Hormone-exposure protocols to trigger the de novo formation of either roots or shoots from callus tissue demonstrate the importance of auxin and cytokinin signaling pathways, and genetic differences in these pathways may contribute to the highly divergent responsiveness of plant species to regeneration protocols. In this study, we show that signaling through MONOPTEROS (MP)/AUXIN RESPONSE FACTOR 5 is necessary for the formation of shoots from Arabidopsis calli. Most strikingly, an irrepressible variant of MP, MPΔ, is sufficient for promoting de novo shoot formation through pathways involving the genetically downstream functions of SHOOT MERISTEMLESS (STM) and CYTOKININ RESPONSE FACTOR2 (CRF2). We conclude that the MPΔ genotype can promote de novo shoot formation and can be used to probe corresponding signaling pathways.

SEUSS and SEUSS-LIKE 2 coordinate auxin distribution and KNOXI activity during embryogenesis.

In Arabidopsis, SEUSS (SEU) and SEUSS-LIKE 2 (SLK2) are components of the LEUNIG (LUG) repressor complex that coordinates various aspects of post-embryonic development. The complex also plays a critical role during embryogenesis, as seu slk2 double mutants have small, narrow cotyledons and lack a shoot apical meristem (SAM). Here we show that seu slk2 double mutant embryos exhibit delayed cotyledon outgrowth and that this is associated with altered PIN-FORMED1 (PIN1) expression and localisation during the early stages of embryogenesis. These observations suggest that SEU and SLK2 promote the transition to bilateral symmetry by modulating auxin distribution in the embryonic shoot. This study also shows that loss of SAM formation in seu slk2 mutants is associated with reduced expression of the class I KNOX (KNOXI) genes SHOOTMERISTEMLESS (STM), BREVIPEDICELLUS and KNAT2. Furthermore, elevating STM expression in seu slk2 mutant embryos was sufficient to restore SAM formation but not post-embryonic activity, while both SAM formation and activity were rescued when SLK2 expression was restored in either the cotyledons or boundary regions. These results demonstrate that SEU and SLK2 function redundantly to promote embryonic shoot development and likely act through a non-cell autonomous pathway to promote KNOXI activity.

Transcriptome profiling reveals roles of meristem regulators and polarity genes during fruit trichome development in cucumber (Cucumis sativus L.).

Trichomes are epidermal hair-like structures that function in plant defence against biotic and abiotic stresses. Extensive studies have been performed on foliar trichomes development in Arabidopsis and tomato, but the molecular mechanism of fruit trichome formation remains elusive. Cucumber fruit is covered with trichomes (spines) that directly affect the appearance and quality of cucumber products. Here, we characterized the fruit spine development in wild-type (WT) cucumber and a spontaneous mutant, tiny branched hair (tbh). Our data showed that the cucumber trichome was multicellular and non-glandular, with malformed organelles and no endoreduplication. Fruit spine development was generally homogenous and marked by a rapid base expansion stage. Trichomes in the tbh mutant were tiny and branched, with increased density and aberrant cell shape. Transcriptome profiling indicated that meristem-related genes were highly enriched in the upregulated genes in the tbh versus the WT, as well as in WT spines after versus before base expansion, and that polarity regulators were greatly induced during spine base expansion. Quantitative reverse transcription PCR and in situ hybridization confirmed the differential expression of CUP-SHAPED COTYLEDON3 (CUC3) and SHOOT MERISTEMLESS (STM) during spine development. Therefore, cucumber trichomes are morphologically different from those of Arabidopsis and tomato, and their development may be regulated by a distinct pathway involving meristem genes and polarity regulators.

The CUC1 and CUC2 genes promote carpel margin meristem formation during Arabidopsis gynoecium development.

Carpel margin meristems (CMMs), a pair of meristematic tissues present along the margins of two fused carpel primordia of Arabidopsis thaliana, are essential for the formation of ovules and the septum, two major internal structures of the gynoecium. Although a number of regulatory factors involved in shoot meristem activity are known to be required for the formation of these gynoecial structures, their direct roles in CMM development have yet to be addressed. Here we show that the CUP-SHAPED COTYLEDON genes CUC1 and CUC2, which are essential for shoot meristem initiation, are also required for formation and stable positioning of the CMMs. Early in CMM formation, CUC1 and CUC2 are also required for expression of the SHOOT MERISTEMLESS gene, a central regulator for stem cell maintenance in the shoot meristem. Moreover, plants carrying miR164-resistant forms of CUC1 and CUC2 resulted in extra CMM activity with altered positioning. Our results thus demonstrate that the two regulatory proteins controlling shoot meristem activity also play critical roles in elaboration of the female reproductive organ through the control of meristematic activity.

STM/BP-Like KNOXI Is Uncoupled from ARP in the Regulation of Compound Leaf Development in Medicago truncatula.

Class I KNOTTED-like homeobox (KNOXI) genes are critical for the maintenance of the shoot apical meristem. The expression domain of KNOXI is regulated by ASYMMETRIC LEAVES1/ROUGHSHEATH2/PHANTASTICA (ARP) genes, which are associated with leaf morphology. In the inverted repeat-lacking clade (IRLC) of Fabaceae, the orthologs of LEAFY (LFY) function in place of KNOXI to regulate compound leaf development. Here, we characterized loss-of-function mutants of ARP (PHAN) and SHOOTMERISTEMLESS (STM)- and BREVIPEDICELLUS (BP)-like KNOXI in the model IRLC legume species Medicago truncatula. The function of ARP genes is species specific. The repression of STM/BP-like KNOXI genes in leaves is not mediated by PHAN, and no suppression of PHAN by STM/BP-like KNOXI genes was observed either, indicating that STM/BP-like KNOXI genes are uncoupled from PHAN in M. truncatula. Furthermore, comparative analyses of phenotypic output in response to ectopic expression of KNOXI and the M. truncatula LFY ortholog, SINGLE LEAFLET1 (SGL1), reveal that KNOXI and SGL1 regulate parallel pathways in leaf development. We propose that SGL1 probably functions in a stage-specific manner in the regulation of the indeterminate state of developing leaves in M. truncatula.

Class I BASIC PENTACYSTEINE factors regulate HOMEOBOX genes involved in meristem size maintenance

The BASIC PENTACYSTEINE (BCP) family is a poorly characterized plant transcription factor family of GAGA BINDING PROTEINS. In Arabidopsis, there are seven members (BPC1-7) that are broadly expressed, and they can potentially bind more than 3000 Arabidopsis GAGA-repeat-containing genes. To date, BPCs are known to be direct regulators of the INNER NO OUTER (INO), SEEDSTICK (STK), and LEAFY COTYLEDON 2 (LEC2) genes. Because of the high functional redundancy, neither single knockout nor double bpc mutant combinations cause aberrant phenotypes. The bpc1-2 bpc2 bpc3 triple mutant shows several pleiotropic developmental defects, including enlargement of the inflorescence meristem and flowers with supernumerary floral organs. Here, we demonstrated through expression analysis and chromatin immunoprecipitation assays that this phenotype is probably due to deregulation of the expression of the SHOOTMERISTEMLESS (STM) and BREVIPEDICELLUS/KNAT1 (BP) genes, which are both direct targets of BPCs. Moreover, we assigned a role to BPCs in the fine regulation of the cytokinin content in the meristem, as both ISOPENTENYLTRANSFERASE 7 (IPT7) and ARABIDOPSIS RESPONSE REGULATOR 7 (ARR7) genes were shown to be overexpressed in the bpc1-2 bpc2 bpc3 triple mutant.

Acquisition and diversification of tendrilled leaves in Bignonieae (Bignoniaceae) involved changes in expression patterns of SHOOTMERISTEMLESS (STM), LEAFY/FLORICAULA (LFY/FLO), and PHANTASTICA (PHAN)

Leaves have undergone structural modifications over evolutionary time, and presently exist in many forms. For instance, in Fabaceae and Bignoniaceae, leaf parts can be modified into tendrils. Currently, no data are available on genic control of tendrilled leaf development outside Fabaceae. Here, we conducted a detailed study of three representatives of Bignonieae: Amphilophium buccinatorium, Dolichandra unguis-cati, and Bignonia callistegioides, bearing multifid, trifid, and simple-tendrilled leaves, respectively. We investigated the structure of their petioles, petiolules, leaflets, and tendrils through histological analyses. Additionally, the expression of SHOOTMERISTEMLESS (STM), PHANTASTICA (PHAN), and LEAFY/FLORICAULA (LFY/FLO) during leaf development was analyzed by in situ hybridizations. Tendrils share some anatomical similarities with leaflets, but not with other leaf parts. Transcripts of both STM and LFY/FLO were detected in leaf primordia, associated with regions from which leaflets and tendril branches originate. PHAN expression was found to be polarized in branched tendrils, but not in simple tendrils. In Bignonieae, tendrils are modified leaflets that, as a result of premature completion of development, become bladeless organs. Bignonieae leaves develop differently from those of peas, as both LFY/FLO and STM are expressed in developing leaves of Bignonieae. Moreover, PHAN is probably involved in tendril diversification in Bignonieae, as it has distinct expression patterns in different leaf types.

Ectopic Expression of BraYAB1-702, a Member of YABBY Gene Family in Chinese Cabbage, Causes Leaf Curling, Inhibition of Development of Shoot Apical Meristem and Flowering Stage Delaying in Arabidopsis thaliana

YABBY gene family plays an important role in the polarity development of lateral organs. We isolated the BraYAB1-702 gene, a member of the YABBY gene family, from young leaves of Chinese cabbage line 06J45. The full-length gene has a 937 bp CDNA sequence and contains an open reading frame (ORF) of 702 bp. The subcellular localization analysis showed that the expression product of the gene was localized in the nucleus. Ectopic expression of BraYAB1-702 in Arabidopsis thaliana caused leaf curling from the adaxial epidermises to abaxial epidermises; the partial abaxialization of the adaxial epidermises of leaves; leaf trichomes and stomata numbers being significantly increased; the plants being severely stunted; the flowering stage being remarkably delayed and inhibiting the development of shoot apical meristem (SAM) with the down-regulation of the expression of SHOOT MERISTEMLESS (STM), Brevipedicellus (BP) and KNAT2 which were related to the development of shoot apical meristem. These results from the present research help to reveal the molecular mechanism of BraYAB1-702 gene in the establishment of adaxial–abaxial polarity of the lateral organs in Chinese cabbage.

The Arabidopsis homeobox gene SHOOT MERISTEMLESS has cellular and meristem‐organisational roles with differential requirements for cytokinin and CYCD3 activity

The Arabidopsis class-1 KNOX gene SHOOT MERISTEMLESS (STM) encodes a homeodomain transcription factor essential for shoot apical meristem (SAM) formation and sustained activity. STM activates cytokinin (CK) biosynthesis in the SAM, but the extent to which STM function is mediated through CK is unclear. Here we show that STM inhibits cellular differentiation and endoreduplication, acting through CK and the CK-inducible CYCD3 cell cycle regulators, establishing a mechanistic link to cell cycle control which provides sustained mitotic activity to maintain a pool of undifferentiated cells in the SAM. Equivalent functions are revealed for the related KNOX genes KNAT1/BP and KNAT2 through ectopic expression. STM is also required for proper meristem organisation and can induce de novo meristem formation when expressed ectopically, even when CK levels are reduced or CK signaling is impaired. This function in meristem establishment and organisation can be replaced by KNAT1/BP, but not KNAT2, despite its activation of CK responses, suggesting that promotion of CK responses alone is insufficient for SAM organisation. We propose that STM has dual cellular and meristem-organisational functions that are differentially represented in the class-1 KNOX gene family and have differing requirements for CK and CYCD3.

Duplicated STM-like KNOX I genes act in floral meristem activity in Eschscholzia californica (Papaveraceae)

In angiosperms, the shoot apical meristem is at the origin of leaves and stems and is eventually transformed into the floral meristem. Class I knotted-like homeobox (KNOX I) genes are known as crucial regulators of shoot meristem formation and maintenance. KNOX I genes maintain the undifferentiated state of the apical meristem and are locally downregulated upon leaf initiation. In Arabidopsis, KNOX I genes, especially SHOOTMERISTEMLESS (STM), have been shown to regulate flower development and the formation of carpels. We investigated the role of STM-like genes in the reproductive development of Eschscholzia californica, to learn more about the evolution of KNOX I gene function in basal eudicots. We identified two orthologs of STM in Eschscholzia, EcSTM1 and EcSTM2, which are predominantly expressed in floral tissues. In contrast, a KNAT1/BP-like and a KNAT2/6-like KNOX I gene are mainly expressed in vegetative organs. Virus-induced gene silencing (VIGS) was used to knockdown gene expression, revealing that both EcSTM genes are required for the formation of reproductive organs. Silencing of EcSTM1 resulted in the loss of the gynoecium and a reduced number of stamens. EcSTM2-VIGS flowers had reduced and defective gynoecia and a stronger reduction in the number of stamen than observed in EcSTM1-VIGS. Co-silencing of both genes led to more pronounced phenotypes. In addition, silencing of EcSTM2 alone or together with EcSTM1 resulted in altered patterns of internodal elongation and sometimes in other floral defects. Our data suggest that some aspects of STM function present in Arabidopsis evolved already before the basal eudicots diverged from core eudicots.

Origin and development of the cryptic shoot meristem in Zeylanidium lichenoides (Podostemaceae)

The shoot apical meristem is the source of aerial shoot systems. In the aquatic eudicot family Podostemaceae, subfamily Podostemoideae, however, shoots develop in the absence of a distinct apical meristem. Previous studies suggest that the cryptic embryonic shoot meristem is involved in primary shoot development in some species (e.g., Zeylanidium lichenoides), although it is unclear whether the meristem maintains meristematic identity. Our aim was to determine how an embryonic shoot meristem is established during embryogenesis and how it is involved in plumular leaf development in the seedling. We observed anatomy and analyzed gene expression in Z. lichenoides, using an ortholog of the shoot meristem marker gene SHOOT MERISTEMLESS (STM). Expression of the STM ortholog, ZlSTM, began in the apical part of the 16-cell embryo. By the heart-shaped embryo stage, its expression was restricted to the putative organizing center (OC) and the protodermal cells just above them, forming a cryptic embryonic shoot meristem without a typical stem cell (apical initials) layer. During seedling development, expression was not maintained in the meristem, but instead shifted to the adaxial bases of cotyledons where plumular leaves would form. ZlSTM expression demonstrated that the meristematic identity is partly retained in the embryonic shoot apex. This cryptic embryonic shoot meristem has a putative OC, but no typical stem cell layer, and it is not maintained during primary shoot development. Modification of the regulatory mechanism between the OC and stem cells might be responsible for this ephemeral shoot meristem in Podostemaceae.

CaHAM is autoregulated and regulates CaSTM expression and is required for shoot apical meristem organization in pepper

The angiosperm shoot apical meristem (SAM) is characterized by tightly organized cell layers and zones. The SAM's organization allows it to maintain its indeterminate nature while producing determinate lateral organs. Alterations in SAM gene expression partly account for the immense diversity in plant architecture. The GRAS protein family gene HAIRY MERISTEM (HAM) is an important regulator of SAM organization in Petunia and Arabidopsis. Here we describe CaHAM loss-of-function pepper mutants characterized by an arrested SAM following the formation of several leaves on the primary stem, complete inhibition of axillary meristem development, an expanded tunica domain and trichome formation on the SAM epidermis. CaHAM is expressed in the periphery of the SAM and in the vasculature of young leaves throughout plant development, reaching its highest level in the reproductive growth stage. Analysis of the effect of CaHAM loss-of-function on its own expression showed that CaHAM is negatively autoregulated. Furthermore, CaHAM negatively regulates the expression level and pattern of pepper SHOOT MERISTEMLESS (CaSTM), which is required to maintain the SAM in an undifferentiated state. We conclude that CaHAM is regulated to achieve adjusted functional levels and has a conserved role in controlling SAM maintenance, organization and axillary meristem formation.