Biology subjects

Immink, R.

Publications and source records attributed to Immink, R..

5 recordsLinked to original sources

bioRxiv2026

An integrative single-cell and spatial transcriptomics atlas highlights candidate regulatory factors in the development of gerbera capitulum

In Asteraceae species, the capitulum is a compact inflorescence, featuring a characteristic reproductive structure. Despite the identification of a few key regulatory factors, the transcriptome-level information on the developing capitulum remains limited. Here, we applied single-cell and spatial transcriptome sequencing to investigate the developing Gerbera hybridas capitulum during floret differentiation. We obtained a transcriptomics atlas encompassing different stages of the Gerbera capitulum and analyzed the cellular and spatial dynamics of gene expression. Using marker gene expression and GO enrichment of cluster-specific DEGs, we annotated putative cell types and described changes in gene expression across sampled stages, potentially associated with ongoing developmental processes. We detected activity of previously undescribed MADS-box genes and defined their spatial expression patterns. Notably, the MADS-box gene GAGL12 was found to be enriched in the putative capitulum phloem cells. The GAGL12 protein was shown in yeast two-hybrid assays to interact with several other MADS-domain proteins with hypothesized functions in vasculature development, and further detailed in silico analyses supported a candidate role in the development of capitulum vasculature. Altogether, we provide integrative and dynamic transcriptomic insight into capitulum and floret development and lay a basis for future functional studies of the control and development of this intriguing reproductive structure.

plant biology↗

bioRxiv2021

The TCP transcription factor HvTB2 heterodimerizes with VRS5(HvTB1) and controls spike architecture in barley

Barley is the fourth largest cereal crop grown worldwide, and essential for food and feed production. Phenotypically, the barley spike, which is unbranched, occurs in two main architectural shapes: two-rowed or six-rowed. In the 6-rowed cultivars, all three florets of the triple floret meristem develop into seeds while in 2-rowed lines only the central floret forms a seed. VRS5(HvTB1), act as inhibitor of lateral seed outgrowth and vrs5(hvtb1) mutants display a six-rowed spike architecture. VRS5(HvTB1) is a member of the TCP transcription factor (TF) family, which often form protein-protein interactions with other transcriptional regulators to modulate the expression of their target genes. Despite the key role of VRS5(HvTB1) in regulating barley plant architecture, there is hardly any knowledge on its molecular mode-of-action. We performed an extensive phylogenetic analysis of the TCP transcription factor family, followed by an in-vitro protein-protein interaction study using yeast-two-hybrid. Our analysis shows that VRS5(HvTB1) has a diverse interaction capacity, interacting with class II TCPs, NF-Y TF, but also chromatin modellers. Further analysis of the interaction capacity of VRS5(HvTB1) with other TCP TFs shows that VRS5(HvTB1) preferably interacts with other class II TCP TFs within the TB1 clade. One of these interactors, encoded by HvTB2, shows a similar expression pattern when compared to VRS5(HvTB1). Haplotype analysis of HvTB2 suggest that this gene is highly conserved and shows hardly any variation in cultivars or wild barley. Induced mutations in HvTB2 trough CRISPR-CAS9 mutagenesis in cv. Golden Promise resulted in barley plants that lost their characteristic unbranched spike architecture. hvtb2 mutants exhibited branches arising at the main spike, suggesting that, similar to VRS5(HvTB1), HvTB2 act as inhibitor of branching. Taken together, our protein-protein interaction studies of VRS5(HvTB1) resulted in the identification of HvTB2, another key regulator of spike architecture in barley. Understanding the molecular network, including protein-protein interactions, of key regulators of plant architecture such as VRS5(HvTB1) provide new routes towards the identification of other key regulators of plant architecture in barley. Author summaryTranscriptional regulation is one of the basic molecular processes that drives plant growth and development. The key TCP transcriptional regulator TEOSINTE BRANCHED 1 (TB1) is one of these key regulators that has been targeted during domestication of several crops for its role as modulator of branching. Also in barley, a key cereal crop, HvTB1 (also referred to as VRS5), inhibits the outgrowth or side shoots, or tillers, and seeds. Despite its key role in barley development, there is hardly any knowledge on the molecular network that is utilized by VRS5(HvTB1). Transcriptional regulators form homo- and heterodimers to regulate the expression of their downstream targets. Here, we performed an extensive phylogenetic analysis of TCP transcription factors (TFs) in barley, followed by protein-protein interaction studies of VRS5(HvTB1). Our analysis indicates, that VRS5(HvTB1) has a diverse capacity of interacting with class II TCPs, NF-Y TF, but also chromatin modellers. Induced mutagenesis trough CRISPR-CAS mutagenesis of one of the putative VRS5(HvTB1) interactors, HvTB2, resulted in barley plants with branched spikes. This shows that insight into the VRS5(HvTB1) interactome, followed by detailed functional analysis of potential interactors is essential to truly understand how TCPs modulate plant architecture. The study presented here provides a first step to underpin the protein-protein interactome of VRS5(HvTB1) and identify other, yet unknown, key regulators of barley plant architecture.

plant biology↗

bioRxiv2021

The Intervening Domain Is Required For DNA-binding and Functional Identity of Plant MADS Transcription Factors

The MADS transcription factors (TF) are an ancient protein family with a high degree of sequence identity that bind almost identical DNA sequences across all eukaryotic kingdoms of life, yet fulfill dramatically different physiological roles. In plants, the family is divided into two main lineages, type I and II, based on sequence conservation of the DNA-binding MADS-box domain (M domain) with yeast and animal M domains. Here, we demonstrate that DNA binding in both lineages absolutely requires a short amino acid sequence C-terminal to the M domain called the Intervening domain (I domain) in type II MADS. Structural elucidation of the MI domains from the floral regulator, SEPALLATA3 (SEP3), shows a highly conserved MADS-box fold with the I domain forming an alpha helix and acting to stabilize the M domain. Based on secondary structure prediction, sequences fulfilling the same function as the SEP3 I domain can be found in both lineages of plant MADS TFs, suggesting the I domain is a conserved and required part of the DNA-binding domain. Using the floral organ identity MADS TFs, SEP3, APETALA1 (AP1) and AGAMOUS (AG), domain swapping demonstrate that the I domain alters DNA-binding specificity based on seq-DAP-seq experiments. Yeast 2-hybrid experiments further revealed the role of the I domain in dimerization specificity. Surprisingly, introducing AG carrying the I domain of AP1 in the Arabidopsis ap1 mutant, resulted in a high degree of complementation and restoration of first and second whorl organs. Taken together, these data demonstrate that the I domain acts both as an integral part of the DNA-binding domain and strongly contributes to the functional identity of the MADS TF.

plant biology↗

bioRxiv2020

A gene regulatory network critical for axillary bud dormancy directly controlled by Arabidopsis BRANCHED1

The control of branch outgrowth is critical for plant fitness, stress resilience and crop yield. The Arabidopsis thaliana transcription factor BRANCHED1 (BRC1) plays a pivotal role in this process as it integrates signals that inhibit axillary bud growth to control shoot branching. Despite the remarkable activity of BRC1 as a potent growth inhibitor, the mechanisms by which it promotes and maintains bud dormancy are still largely unknown. Here we combine ChIP-seq, transcriptomic and systems biology approaches to characterise the BRC1-regulated gene network. We identify a group of BRC1 direct target genes encoding transcription factors (BTFs) that orchestrate, together with BRC1, an intricate transcriptional network enriched in abscisic acid signalling components. The BRC1 network is enriched in feed-forward loops and feed-back loops, robust against noise and mutation, reversible in response to stimuli, and stable once established. This knowledge is fundamental to adapt plant architecture and crop production to ever-changing environmental conditions.

plant biology↗

bioRxiv2018

A molecular framework for functional versatility of HECATE transcription factors

During the plant life cycle, diverse signalling inputs are continuously integrated and engage specific genetic programs depending on the cellular or developmental context. Consistent with an important role in this process, HECATE (HEC) bHLH transcription factors display diverse functions, from photomorphogenesis to the control of shoot meristem dynamics and gynoecium patterning. However, the molecular mechanisms underlying their functional versatility and the deployment of specific HEC sub-programs still remain elusive.\n\nTo address this issue, we systematically identified proteins with the capacity to interact with HEC1, the best characterized member of the family, and integrated this information with our data set of direct HEC1 target genes. The resulting core genetic modules were consistent with specific developmental functions of HEC1, including its described activities in light signalling, gynoecium development and auxin homeostasis. Importantly, we found that in addition, HEC genes play a role in the modulation of flowering time and uncovered that their role in gynoecium development may involve the direct transcriptional regulation of NGATHA1 (NGA1) and NGA2 genes. NGA factors were previously shown to contribute to fruit development, but our data now show that they also modulate stem cell homeostasis in the SAM.\n\nTaken together, our results suggest a molecular network underlying the functional versatility of HEC transcription factors. Our analyses have not only allowed us to identify relevant target genes controlling shoot stem cell activity and a so far undescribed biological function of HEC1, but also provide a rich resource for the mechanistic elucidation of further context dependent HEC activities.\n\nSignificance statementAlthough many transcription factors display diverse regulatory functions during plant development, our understanding of the underlying mechanisms remains poor. Here, by reconstructing the regulatory modules orchestrated by the bHLH transcription factor HECATE1 (HEC1), we defined its regulatory signatures and delineated a regulatory network that provides a molecular basis for its functional versatility. In addition, we uncovered a function for HEC genes in modulating flowering time and further identified downstream signalling components balancing shoot stem cell activity.

plant biology↗