HAM Gene Family and Shoot Meristem Development

Land plants develop highly diversified shoot architectures, all of which are derived from the pluripotent stem cells in shoot apical meristems (SAMs). As sustainable resources for continuous organ formation in the aboveground tissues, SAMs play an important role in determining plant yield and biomass production. In this review, we summarize recent advances in understanding one group of key regulators – the HAIRY MERISTEM (HAM) family GRAS domain proteins – in shoot meristems. We highlight the functions of HAM family members in dictating shoot stem cell initiation and proliferation, the signaling cascade that shapes HAM expression domains in shoot meristems, and the conservation and diversification of HAM family members in land plants. We also discuss future directions that potentially lead to a more comprehensive view of the HAM gene family and stem cell homeostasis in land plants.

Distinct within-host bacterial populations ensure function, colonization and transmission in leaf symbiosis

Hereditary symbioses have the potential to drive transgenerational effects, yet the mechanisms responsible for transmission of heritable plant symbionts are still poorly understood. The leaf symbiosis between Dioscorea sansibarensis and the bacterium Orrella dioscoreae offers an appealing model system to study how heritable bacteria are transmitted to the next generation. Here, we demonstrate that inoculation of apical buds with a bacterial suspension is sufficient to colonize newly-formed leaves and propagules, and to ensure transmission to the next plant generation. Flagellar motility is not required for movement inside the plant, but is important for the colonization of new hosts. Further, stringent tissue-specific regulation of putative symbiotic functions highlight the presence of two distinct subpopulations of bacteria in the leaf gland and at the shoot meristem. We propose that bacteria in the leaf gland dedicate resources to symbiotic functions, while dividing bacteria in the shoot tip ensure successful colonization of meristematic tissue, glands and propagules. Compartmentalization of intra-host populations, together with tissue-specific regulation may serve as a robust mechanism for the maintenance of mutualism in leaf symbiosis.ImportanceSeveral plant species form associations with bacteria in their leaves, called leaf symbiosis. These associations are highly specific, but the mechanisms responsible for symbiont transmission are poorly understood. Using the association between the yam species Dioscorea sansibarensis and Orrella dioscoreae as a model leaf symbiosis, we provide experimental evidence that bacteria are transmitted vertically and distributed to specific leaf structures via association with shoot meristems. Flagellar motility is required for initial infection, but does not contribute to spread within host tissue. We also provide evidence that bacterial subpopulations at the meristem or in the symbiotic leaf gland differentially express key symbiotic genes. We argue that this separation of functional symbiont populations, coupled to tight control over bacterial infection and transmission, explain the evolutionary robustness of leaf symbiosis. These findings may provide insights into how plants may recruit and maintain beneficial symbionts at the leaf surface.

Regulatory circuits involving bud dormancy factor PpeDAM6

DORMANCY-ASSOCIATED MADS-BOX (DAM) genes have recently emerged as key potential regulators of the dormancy cycle and climate adaptation in perennial species. Particularly, PpeDAM6 has been proposed to act as a major repressor of bud dormancy release and bud break in peach (Prunus persica). PpeDAM6 expression is downregulated concomitantly with the perception of a given genotype-dependent accumulation of winter chilling time, and the coincident enrichment in H3K27me3 chromatin modification at a specific genomic region. We have identified three peach BASIC PENTACYSTEINE PROTEINs (PpeBPCs) interacting with two GA-repeat motifs present in this H3K27me3-enriched region. Moreover, PpeBPC1 represses PpeDAM6 promoter activity by transient expression experiments. On the other hand, the heterologous overexpression of PpeDAM6 in European plum (Prunus domestica) alters plant vegetative growth, resulting in dwarf plants tending toward shoot meristem collapse. These alterations in vegetative growth of transgenic lines associate with impaired hormone homeostasis due to the modulation of genes involved in jasmonic acid, cytokinin, abscisic acid, and gibberellin pathways, and the downregulation of shoot meristem factors, specifically in transgenic leaf and apical tissues. The expression of many of these genes is also modified in flower buds of peach concomitantly with PpeDAM6 downregulation, which suggests a role of hormone homeostasis mechanisms in PpeDAM6-dependent maintenance of floral bud dormancy and growth repression.

Detection of Target Sites Edited in Upland Cotton By The CRISPR/Cas9 System Mediated By Agrobacterium Rhizogenes

Background CRIPSR/Cas9 gene editing has the ability to effectively modify plant genomes. Multiple target sites usually were designed and the effective target sites were selected for editing. However, upland cotton is allotetraploid and is commonly considered as difficult and inefficient to transform. Therefore, it’s important to quickly identify feasibility of the target site. In this study, we use Agrobacterium rhizogenes K599 strain to infect cotton shoot meristem and induce them to grow hairy roots to detect the feasibility of a selected target designed in GhMYB25-like gene. Results We designed a sgRNA within the second exons of GhMYB25-likeA and GhMYB25-likeD and constructed the CRISPR vector. Transient hairy root transformation using A. rhizogenes K599 with four OD600s (0.4, 0.6,0.8, 1.0) was performed in Coker 312 (R15). The results show that A. rhizogenes at OD600 = 0.6–0.8 is the best concentration range for inducing cotton hairy roots. The other three cultivars (TM-1, Lumian 21, Zhongmian 49) were injected using A. rhizogenes K599 with OD600 = 0.6-0.8 and all produced hairy roots. We characterized ten R15 plants with hairy roots and detected different degrees of base deletions and insert at the target site in five R15 plants. Conclusion Overall, our data show A. rhizogenes-mediated transient hairy root transformation offers a rapid and efficient method to detect sgRNA feasibility in cotton.

Genetics of Shoot Meristem and Shoot Regeneration

Plants exhibit remarkable lineage plasticity, allowing them to regenerate organs that differ from their respective origins. Such developmental plasticity is dependent on the activity of pluripotent founder cells or stem cells residing in meristems. At the shoot apical meristem (SAM), the constant flow of cells requires continuing cell specification governed by a complex genetic network, with the WUSCHEL transcription factor and phytohormone cytokinin at its core. In this review, I discuss some intriguing recent discoveries that expose new principles and mechanisms of patterning and cell specification acting both at the SAM and, prior to meristem organogenesis during shoot regeneration. I also highlight unanswered questions and future challenges in the study of SAM and meristem regeneration. Finally, I put forward a model describing stochastic events mediated by epigenetic factors to explain how the gene regulatory network might be initiated at the onset of shoot regeneration. Expected final online publication date for the Annual Review of Genetics, Volume 55 is November 2021. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.

Asymmetric expansions of FT and TFL1 lineages characterize differential evolution of the EuPEBP family in the major angiosperm lineages

Background In flowering plants, precise timing of the floral transition is crucial to maximize chances of reproductive success, and as such, this process has been intensively studied. FLOWERING LOCUS T (FT) and TERMINAL FLOWER1 (TFL1) have been identified as closely related eukaryotic phosphatidylethanolamine-binding proteins (‘EuPEBPs’) that integrate multiple environmental stimuli, and act antagonistically to determine the optimal timing of the floral transition. Extensive research has demonstrated that FT acts similar to hormonal signals, being transported in the phloem from its primary site of expression in leaves to its primary site of action in the shoot meristem; TFL1 also appears to act as a mobile signal. Recent work implicates FT, TFL1, and the other members of the EuPEBP family, in the control of other important processes, suggesting that the EuPEBP family may be key general regulators of developmental transitions in flowering plants. In eudicots, there are a small number of EuPEBP proteins, but in monocots, and particularly grasses, there has been a large, but uncharacterized expansion of EuPEBP copy number, with unknown consequences for the EuPEBP function. Results To systematically characterize the evolution of EuPEBP proteins in flowering plants, and in land plants more generally, we performed a high-resolution phylogenetic analysis of 701 PEBP sequences from 208 species. We refine previous models of EuPEBP evolution in early land plants, demonstrating the algal origin of the family, and pin-pointing the origin of the FT/TFL1 clade at the base of monilophytes. We demonstrate how a core set of genes (MFT1, MFT2, FT, and TCB) at the base of flowering plants has undergone differential evolution in the major angiosperm lineages. This includes the radical expansion of the FT family in monocots into 5 core lineages, further re-duplicated in the grass family to 12 conserved clades. Conclusions We show that many grass FT proteins are strongly divergent from other FTs and are likely neo-functional regulators of development. Our analysis shows that monocots and eudicots have strongly divergent patterns of EuPEBP evolution.

Genome-Wide Identification and Characterization of KNOTTED-Like Homeobox (KNOX) Homologs in Garlic (Allium sativum L.) and Their Expression Profilings Responding to Exogenous Cytokinin and Gibberellin

Garlic (Allium sativum L.) is an important vegetable and is cultivated and consumed worldwide for its economic and medicinal values. Garlic cloves, the major reproductive and edible organs, are derived from the axillary meristems. KNOTTED-like homeobox (KNOX) proteins, such as SHOOT MERISTEM-LESS (STM), play important roles in axillary meristem formation and development. However, the KNOX proteins in garlic are still poorly known. Here, 10 AsKNOX genes, scattered on 5 of the 8 chromosomes, were genome-wide identified and characterized based on the newly released garlic genome. The typical conserved domains of KNOX proteins were owned by all these 10 AsKNOX homologs, which were divided into two Classes (Class I and Class II) based on the phylogenetic analysis. Prediction and verification of the subcellular localizations revealed the diverse subcellular localization of these 10 AsKNOX proteins. Cis-element prediction, tissue expression analysis, and expression profilings in responding to exogenous GA3 and 6-BA showed the potential involvement of AsKNOX genes in the gibberellin and cytokinin signaling pathways. Overall, the results of this work provided a better understanding of AsKNOX genes in garlic and laid an important foundation for their further functional studies.

Cytokinin-CLAVATA crosstalk is an ancient mechanism regulating shoot meristem homeostasis in land plants

Plant shoots grow from stem cells within Shoot Apical Meristems (SAMs), which produce lateral organs while maintaining the stem cell pool. In the model flowering plant Arabidopsis, the CLAVATA (CLV) pathway functions antagonistically with cytokinin signaling to control the size of the multicellular SAM via negative regulation of the stem cell organizer WUSCHEL (WUS). Although comprising just a single cell, the SAM of the model moss Physcomitrium patens (formerly Physcomitrella) performs equivalent functions during stem cell maintenance and organogenesis, despite the absence of WUS-mediated stem cell organization. Our previous work showed that the stem cell-delimiting function of the CLV pathway receptors CLAVATA1 (CLV1) and RECEPTOR-LIKE PROTEIN KINASE2 (RPK2) is conserved in the moss P. patens. Here, we use P. patens to assess whether CLV-cytokinin crosstalk is also an evolutionarily conserved feature of stem cell regulation. Genetic analyses reveal that CLV1 and RPK2 regulate SAM proliferation via separate pathways in moss. Surprisingly, cytokinin receptor mutants also form ectopic stem cells in the absence of cytokinin signaling. Through modeling, we identified regulatory network archtectures that recapitulated the stem cell phenotypes of clv1 and rpk2 mutants, cytokinin application, cytokinin receptor mutations, and higher-order combinations of these perturbations. These models predict that CLV1 and RPK2 act through separate pathways wherein CLV1 represses cytokinin-mediated stem cell initiation and RPK2 inhibits this process via a separate, cytokinin-independent pathway. Our analysis suggests that crosstalk between CLV1 and cytokinin signaling is an evolutionarily conserved feature of SAM homeostasis that preceded the role of WUS in stem cell organization.