Biology subjects

Sidoli, S.

Publications and source records attributed to Sidoli, S..

At least 73 records · Page 4Linked to original sources

bioRxiv2022

Proximity labeling reveals a new in vivo network of interactors for the histone demethylase KDM5

BackgroundKDM5 family proteins are multi-domain regulators of transcription that when dysregulated contribute to cancer and intellectual disability. KDM5 proteins can regulate transcription through their histone demethylase activity in addition to demethylase-independent gene regulatory functions that remain less characterized. To expand our understanding of the mechanisms that contribute to KDM5-mediated transcription regulation, we used TurboID proximity labeling to identify KDM5-interacting proteins. ResultsUsing Drosophila melanogaster, we enriched for biotinylated proteins from KDM5-TurboID-expressing adult heads using a newly generated control for DNA-adjacent background in the form of dCas9:TurboID. Mass spectrometry analyses of biotinylated proteins identified both known and novel candidate KDM5 interactors, including members of the SWI/SNF and NURF chromatin remodeling complexes, the NSL complex, Mediator, and several insulator proteins. ConclusionsCombined, our data shed new light on potential demethylase-independent activities of KDM5. In the context of KDM5 dysregulation, these interactions may play key roles in the alteration of evolutionarily conserved transcriptional programs implicated in human disorders.

genetics↗

bioRxiv2022

In vivo proximity ligation reveals endogenous candidate interactors of Neurexin's intracellular domain

Neurexins are highly-spliced transmembrane cell adhesion molecules that bind an array of partners via their extracellular domains. However, much less is known about the signaling pathways downstream of neurexins largely-invariant intracellular domain. C. elegans contains a single neurexin gene that we have previously shown is required for presynaptic assembly and stabilization. To gain insight into the signaling pathways mediating neurexins presynaptic functions, we employed a proximity ligation method, endogenously tagging neurexins intracellular domain with the promiscuous biotin ligase TurboID, allowing us to isolate adjacent biotinylated proteins by streptavidin pull-down and mass spectrometry. We compared our experimental strain to a control strain in which neurexin, endogenously tagged with TurboID, was dispersed from presynaptic active zones by the deletion of its C-terminal PDZ-binding motif. Using this approach we identified both known and novel intracellular interactors of neurexin, including active zone scaffolds, actin-binding proteins (including almost every member of the Arp2/3 complex), signaling molecules, and mediators of RNA trafficking, protein synthesis and degradation, among others. Characterization of mutants for candidate neurexin interactors revealed that they recapitulate aspects of the nrx-1 mutant phenotype, suggesting they may be involved in neurexin signaling. Finally, to investigate a possible role for neurexin in local actin assembly, we endogenously tagged its intracellular domain with actin depolymerizing and sequestering peptides (DeActs), and found that this led to defects in active zone assembly.

neuroscience↗

bioRxiv2022

Investigation of reversible histone acetylation and dynamics in gene expression regulation using 3D liver spheroid model

BackgroundThree-dimensional (3D) cell culture has emerged as an alternative approach to 2D flat culture to model more accurately the phenotype of solid tissue in laboratories. Culturing cells in 3D more precisely recapitulates physiological conditions of tissues, as these cells reduce activities related to proliferation, focusing their energy consumption towards metabolism and homeostasis. ResultsHere, we demonstrate that 3D liver spheroids are a suitable system to model chromatin dynamics and response to epigenetics inhibitors. To delay necrotic tissue formation despite proliferation arrest, we utilize rotating bioreactors that apply active media diffusion and low shearing forces. We demonstrate that the proteome and the metabolome of our model resemble typical liver functions. We prove that spheroids respond to sodium butyrate (NaBut) treatment, an inhibitor of histone deacetylases (HDACi), by upregulating histone acetylations and transcriptional activation. As expected, NaBut treatment impaired specific cellular functions, including the energy metabolism. More importantly, we demonstrate that spheroids reestablish their original proteome and transcriptome, including pre-treatment levels of histone acetylation, metabolism, and protein expression once the standard culture condition is restored after treatment. Given the slow replication rate (>40 days) of cells in 3D spheroids, our model enables to monitor the recovery of approximately the same cells that underwent treatment, demonstrating that NaBut does not have long-lasting effects on histone acetylation and gene expression. These results suggest that histone acetylation has minimal epigenetics memory in our spheroids culture. ConclusionTogether, we established an innovative cell culture system that can be used to model anomalously decondensing chromatin in physiological cell growth and rule out epigenetics inheritance if cells recover the original phenotype after treatment. The transient epigenetics effects demonstrated here highlights the relevance of using a 3D culture model system that could be very useful in studies requiring long term drug treatment conditions that would not be possible using a 2D cell monolayer system.

cell biology↗

bioRxiv2022

Locus-specific proteomics identifies new aspects of the chromatin context involved in V region somatic hypermutation

Activation-induced cytidine deaminase (AID) somatically hypermutates the immunoglobulin heavy chain variable region (IGHV) gene to create the antibody diversity required to resist infections. This hypermutational process involves many pathways including transcription, DNA structural change and repair. While many of the proteins involved have been identified, their relative abundance, organization and regulation have not been resolved and additional factors and pathways need to be identified. To identify the proteome occupying IGHV, we have utilized dCas9-APEX targeted by guide RNAs to biotinylate and enrich the proteins associated with the mutating V region chromatin in the Ramos human B cell line and compared them to the non-mutating downstream constant region (C) chromatin. We identified hundreds of proteins specifically enriched on the V or C region. We confirmed the functionality of selected factors by examining the changes in the V region-specific proteome after inhibiting transcriptional elongation and somatic mutation with the Dot1L inhibitor EPZ004777. SummaryLocus-specific proteomics using dCas9-APEX identifies new aspects of the chromatin context involved in V region somatic hypermutation (SHM) in the human Ramos B cell line. An inhibitor of Dot1L which participates in SHM is used to identify functional SHM-related factors.

immunology↗

bioRxiv2022

mTORC2 couples fasting to mitochondrial fission

Fasting triggers diverse cellular and metabolic adaptations to facilitate organismal survival1,2. During nutrient deprivation, increases in circulating fatty acids support mitochondrial respiration2. The mechanisms driving mitochondrial adaptations and respiratory sufficiency during nutrient deprivation remain incompletely understood. Here we show that extended periods of fasting, or lipid availability stimulates mTORC2 activity. Activation of mTORC2 and phosphorylation of its target NDRG13 at S336 sustains mitochondrial fission and respiratory sufficiency. Timelapse imaging reveals that wildtype NDRG1, but not phosphorylation-deficient NDRG1S336A mutant, engages with mitochondria to facilitate its scission. Using proteomics, and an siRNA screen, we show that mTORC2-phosphorylated NDRG1 cooperates with the small GTPase Cdc424 and Cdc42-specific effectors and regulators to orchestrate fission. Accordingly, RictorKO, NDRG1S336A mutants, and Cdc42-deficient cells each display mitochondrial phenotypes reminiscent of fission failure. During nutrient surplus, mTOR complexes perform anabolic functions5; however, paradoxical reactivation of mTORC2 during fasting plays an unexpected role in driving mitochondrial fission and respiration.

cell biology↗

bioRxiv2022

Cell-Type-Specific Epigenetic Priming of Gene Expression in Nucleus Accumbens by Cocaine

A hallmark of addiction is the ability of drugs of abuse to trigger relapse after periods of prolonged abstinence. Here, we describe a novel epigenetic mechanism whereby chronic cocaine exposure causes lasting chromatin and downstream transcriptional modifications in the nucleus accumbens (NAc), a critical brain region controlling motivation. We link prolonged withdrawal from cocaine to the depletion of the histone variant H2A.Z, coupled to increased genome accessibility and latent priming of gene transcription, in D1 dopamine receptor-expressing medium spiny neurons (D1 MSNs) that relates to aberrant gene expression upon drug relapse. The histone chaperone ANP32E removes H2A.Z from chromatin, and we demonstrate that D1 MSN-selective Anp32e knockdown prevents cocaine-induced H2A.Z depletion and blocks cocaines rewarding actions. By contrast, very different effects of cocaine exposure, withdrawal, and relapse were found for D2-MSNs. These findings establish histone variant exchange as an important mechanism and clinical target engaged by drugs of abuse to corrupt brain function and behavior.

molecular biology↗

bioRxiv2022

TMEM106B is increased in Multiple Sclerosis plaques, and deletion causes accumulation of lipid after demyelination

During inflammatory, demyelinating diseases such as multiple sclerosis (MS) axonal damage is prevalent early in the disease course. Axonal damage includes swellings, defects in transport, and failure to clear damaged intracellular proteins, all of which affect recovery and compromise the integrity of neurons and remyelination. Autophagy and the clearance of damaged cell components by the proteasome are important for the maintenance of normal cellular turnover; and the restoration of cellular homeostasis. The gradual accumulation of insoluble proteins in the brain is known to impair recovery from several neurodegenerative diseases. In this study, we used mass spectrometry to identify insoluble proteins within high-speed, mercaptoethanol/sarcosyl-insoluble pellets from purified white matter plaques isolated from the brains of individuals with MS. We determined that insoluble transmembrane protein106B (TMEM106B), expressed in neurons and glia and normally lysosomal-associated, is increased in MS plaques relative to normal-appearing white matter from individuals with Alzheimers disease and non-neurologic controls. We found that decreased TMEM106B protein in mice results in significant axonal damage and lipid droplet accumulation in the spinal cord following chronic myelin oligodendrocyte glycoprotein-induced experimental autoimmune encephalomyelitis. When TMEM106Bt/t mice were treated with cuprizone to experimentally induce demyelination, a significant increase in lipid deposition was observed in the corpus callosum of TMEM106Bt/t mice post-cuprizone withdrawal. Our study shows that the brain and spinal cord from challenged TMEM106Bt/t mice accumulate OilRedO+/Perilipin2+ lipid droplets. We postulate that increased insolubility of TMEM106B in MS plaques limits debris clearance by the lysosome which over time contributes to failed remyelination and axonal defects. Abbreviated AbstractTransmembrane protein106B (TMEM106B), a lysosome-associated protein, is significantly less soluble in multiple sclerosis plaques than in white matter controls. Decreased TMEM106B produces significant axonal damage and lipid accumulation in mouse models of demyelinating diseases. TMEM106B insolubility and likely loss of function may limit lysosome transport and contribute to CNS pathology.

neuroscience↗

bioRxiv2022

Autophagic state prospectively identifies facultative stem cells in the intestinal epithelium

The intestinal epithelium exhibits a rapid and efficient regenerative response to injury. Emerging evidence supports a model where plasticity of differentiated cells, particularly those in the secretory lineages, contributes to epithelial regeneration upon ablation of injury-sensitive stem cells. However, such facultative stem cell activity is rare within secretory populations. Here we ask whether specific functional properties predict facultative stem cell activity. We utilize in vivo labeling combined with ex vivo organoid formation assays to evaluate how cell age and autophagic state contribute to facultative stem cell activity within secretory lineages. Strikingly, we find that cell age (time elapsed since cell cycle exit) does not correlate with secretory cell plasticity. Instead, high autophagic activity predicts plasticity and resistance to DNA damaging injury independently of cell lineage. Our findings indicate that autophagic status prior to injury serves as a lineageagnostic marker for the prospective identification of facultative stem cells.

cell biology↗

bioRxiv2022

Functional Contribution of Cancer-Associated Fibroblasts in Glioblastoma

The abundance and biological contribution of cancer associated fibroblasts (CAFs) in glioblastoma are poorly understood. Here, we applied single-cell RNA sequencing and spatial transcriptomics analyses to identify and characterize CAFs in human glioblastoma tumors and then performed functional enrichment analysis and in vitro assays to investigate their interactions with malignant glioblastoma cells. We found that CAF abundance was significantly correlated with tumor grade, poor clinical outcome, and activation of extracellular matrix remodeling, using three large databases containing bulk RNA-sequencing data and clinical information. Proteomic analysis of the CAFs and their secretome revealed fibronectin (FN1) as a strong candidate mediating CAF functions. This was validated using in vitro cellular models, which demonstrated that CAF conditioned media and recombinant FN1 could facilitate the migration and invasion of glioblastoma cells. In addition, we showed that CAFs were more abundant in the mesenchymal-like state (or subtype) than in other states of glioblastomas, while cell lines resembling the proneural-state responded to the CAF signaling better in terms of the migratory and invasive phenotypes. Investigating the in-situ expression of gene markers specifically associated with CAFs and mesenchymal malignant cells further indicated that CAFs were enriched in the perinecrotic and pseudopalisading zones of human tumors, where mesenchymal-like glioblastoma cells co-resided and thus likely interacted. Overall, this study characterized the molecular features and functional impacts of CAFs in glioblastoma, alluding to a novel cell-to-cell interaction axis mediated by CAFs in the glioblastoma microenvironment.

cancer biology↗

bioRxiv2022

Epigenetic dysregulation from chromosomal transit in micronuclei

Chromosomal instability (CIN) and epigenetic alterations are characteristics of advanced and metastatic cancers [1-4], yet whether they are mechanistically linked is unknown. Here we show that missegregation of mitotic chromosomes, their sequestration in micronuclei [5, 6], and subsequent micronuclear envelope rupture [7] profoundly disrupt normal histone post-translational modifications (PTMs), a phenomenon conserved across humans and mice as well as cancer and non-transformed cells. Some of the changes to histone PTMs occur due to micronuclear envelope rupture whereas others are inherited from mitotic abnormalities prior to micronucleus formation. Using orthogonal techniques, we show that micronuclei exhibit extensive differences in chromatin accessibility with a strong positional bias between promoters and distal or intergenic regions. Finally, we show that inducing CIN engenders widespread epigenetic dysregulation and that chromosomes which transit in micronuclei experience durable abnormalities in their accessibility long after they have been reincorporated into the primary nucleus. Thus, in addition to genomic copy number alterations, CIN can serve as a vehicle for epigenetic reprogramming and heterogeneity in cancer.

cancer biology↗

bioRxiv2021

Dense granule protein, GRA64 interacts with host cell ESCRT proteins during Toxoplasma gondii infection

The intracellular parasite Toxoplasma gondii adapts to diverse host cell environments within a replicative compartment that is heavily decorated by secreted proteins. In attempts to identify novel parasite secreted proteins that influence host cell activity, we identified and characterized a trans-membrane dense granule protein dubbed GRA64 (TGME49_202620). We found that GRA64 is on the parasitophorous vacuolar membrane (PVM) and is partially exposed to the host cell cytoplasm in both tachyzoite and bradyzoite parasitophorous vacuoles. Using co-immunoprecipitation and proximity-based biotinylation approaches, we demonstrate that GRA64 appears to interact with certain components of the host Endosomal Sorting Complexes Required for Transport (ESCRT). Genetic disruption of GRA64 does not affect acute Toxoplasma virulence in mice nor encystation as observed via tissue cyst burdens in mice during chronic infection. However, ultrastructural analysis of {Delta}gra64 tissue cysts using electron tomography revealed enlarged vesicular structures underneath the cyst membrane, suggesting a role for GRA64 in organizing the recruitment of ESCRT proteins and subsequent intracystic vesicle formation. This study uncovers a novel host-parasite interaction that contributes to an emerging paradigm in which specific host ESCRT proteins are recruited to the limiting membranes (PVMs) of tachyzoite and bradyzoite vacuoles formed during acute and chronic Toxoplasma infection. IMPORTANCEToxoplasma gondii is a widespread foodborne parasite that causes congenital disease and life-threatening complications in immune compromised individuals. Part of this parasites success lies in its ability to infect diverse organisms and host cells, as well as to persist as a latent infection within parasite constructed structures called tissue cysts. In this study, we characterized a protein secreted by T. gondii into its parasitophorous vacuole during intracellular infection, which we dub GRA64. On the vacuole, this protein is exposed to the host cell and interacts with specific host ESCRT proteins. Parasites lacking the GRA64 protein exhibit ultrastructural changes in tissue cysts during chronic infection. This study lays the foundation for future studies on the mechanics and consequences of host ESCRT-parasite protein interactions.

microbiology↗

bioRxiv2021

Multi-omic profiling of histone variant H3.3 lysine 27 methylation reveals a distinct role from canonical H3 in stem cell differentiation

Histone variants, such as histone H3.3, replace canonical histones within the nucleosome to alter chromatin accessibility and gene expression. Although the biological roles of selected histone post-translational modifications (PTMs) have been extensively characterized, the potential differences in the function of a given PTM on different histone variants is almost always elusive. By applying proteomics and genomics techniques, we investigate the role of lysine 27 tri-methylation specifically on the histone variant H3.3 (H3.3K27me3) in the context of mouse embryonic stem cell pluripotency and differentiation as a model system for development. We demonstrate that while the steady state overall levels of methylation on both H3K27 and H3.3K27 decrease during differentiation, methylation dynamics studies indicate that methylation on H3.3K27 is maintained more than on H3K27. Using a custom-made antibody, we identify a unique enrichment of H3.3K27me3 at lineage-specific genes, such as olfactory receptor genes, and at binding motifs for the transcription factors FOXJ2/3. REST, a predicted FOXJ2/3 target that acts as a transcriptional repressor of terminal neuronal genes, was identified with H3.3K27me3 at its promoter region. H3.3K27A mutant cells confirmed an upregulation of FOXJ2/3 targets upon the loss of methylation at H3.3K27. Thus, while canonical H3K27me3 has been characterized to regulate the expression of transcription factors that play a general role in differentiation, our work suggests H3.3K27me3 is essential for regulating distinct terminal differentiation genes. This work highlights the importance of understanding the effects of PTMs not only on canonical histones but also on specific histone variants, as they may exhibit distinct roles.

genomics↗

bioRxiv2021

Genotype-specific Features Reduce the Susceptibility of South American Yellow Fever Virus Strains to Vaccine-Induced Antibodies

The resurgence of yellow fever in South America has prompted mitigation through vaccination against the etiologic agent, yellow fever virus (YFV). Current vaccines are based on a virulent African isolate, and their capacity to induce neutralizing antibodies against the vaccine strain is widely used as a surrogate for protection. However, the sensitivity of genetically distinct South American strains to vaccine-induced antibodies is unknown. Here, we show that antiviral potency of the polyclonal antibody response in both U.S. and Brazilian vaccinees is attenuated against an emergent Brazilian strain. This reduction was attributable to genetic changes at two sites in the central domain II of the glycoprotein E, including the acquisition of an N-linked glycosylation site, which are unique to and shared among most South American YFV strains. Our findings call for a reevaluation of current approaches to YFV immunological surveillance in South America and suggest approaches for designing updated vaccines.

microbiology↗

bioRxiv2021

Type I PRMTs and PRMT5 Inversely Regulate Post-Transcriptional Intron Detention

Protein arginine methyltransferases (PRMTs) are required for the regulation of RNA processing factors. Type I enzymes catalyze mono- and asymmetric dimethylation; Type II enzymes catalyze mono- and symmetric dimethylation. To understand the specific mechanisms of PRMT activity in splicing regulation, we inhibited Type I and II PRMTs and probed their transcriptomic consequences. Using the newly developed SKaTER-seq method, analysis of co-transcriptional splicing revealed that PRMT inhibition resulted in slower splicing rates. Surprisingly, altered co-transcriptional splicing kinetics correlated poorly with ultimate changes in alternative splicing of polyadenylated RNA--particularly intron retention (RI). Investigation of RI following inhibition of nascent transcription demonstrated that PRMTs inversely regulate RI post-transcriptionally. Subsequent proteomic analysis of chromatin-associated polyadenylated RNA identified aberrant binding of the Type I substrate, CHTOP, and the Type II substrate, SmB. Targeted mutagenesis of all methylarginine sites in SmD3, SmB, and SmD1 recapitulated splicing changes seen with Type II PRMT inhibition. Conversely, mutagenesis of all methylarginine sites in CHTOP recapitulated the splicing changes seen with Type I PRMT inhibition. Closer examination of subcellular fractions indicated that RI were isolated to the nucleoplasm and chromatin. Together, these data demonstrate that PRMTs regulate the post-transcriptional processing of nuclear, detained introns through Sm and CHTOP arginine methylation.

cancer biology↗

bioRxiv2021

Cleavage of histone H2A during embryonic stem cell differentiation destabilizes nucleosomes to counteract gene activation

Histone proteolysis is a poorly understood phenomenon in which the N-terminal tails of histones are irreversibly cleaved by intracellular proteases. During development, histone post-translational modifications are known to orchestrate gene expression patterns that ultimately drive cell fate decisions. Therefore, deciphering the mechanisms of histone proteolysis is necessary to enhance the understanding of cellular differentiation. Here we show that H2A is cleaved by the lysosomal protease Cathepsin L during ESCs differentiation. Using quantitative mass spectrometry (MS), we identified L23 to be the primary cleavage site that gives rise to the clipped form of H2A (cH2A), which reaches a maximum level of ~1% of total H2A after four days of differentiation. Using ChIP-seq, we found that preventing proteolysis leads to an increase in acetylated H2A at promoter regions in differentiated ES cells. We also identified novel readers of different acetylated forms of H2A in pluripotent ES cells, such as members of the PBAF remodeling complex. Finally, we showed that H2A proteolysis abolishes this recognition. Altogether, our data suggests that proteolysis serves as an efficient mechanism to silence pluripotency genes and destabilize the nucleosome core particle.

developmental biology↗

bioRxiv2021

Cytoplasmic labile iron accumulates in aging stem cells perturbing a key rheostat for identity control

Bone marrow resident and rarely dividing haematopoietic stem cells (HSC) harbour an extensive self-renewal capacity to sustain life-long blood formation;1-5 albeit their function declines during ageing.6, 7 Various molecular mechanisms confer stem cell identity, ensure long-term maintenance and are known to be deregulated in aged stem cells.8, 9 How these programs are coordinated, particularly during cell division, and what triggers their ageing-associated dysfunction has been unknown. Here, we demonstrate that HSC, containing the lowest amount of cytoplasmic chelatable iron (labile iron pool)10 among hematopoietic cells, activate a limited iron response during mitosis. Engagement of this iron homeostasis pathway elicits mobilization and {beta}-oxidation of arachidonic acid and enhances stem cell-defining transcriptional programs governed by histone acetyl transferase Tip60/KAT5. We further find an age-associated expansion of the labile iron pool, along with loss of Tip60/KAT5-dependent gene regulation to contribute to the functional decline of ageing HSC, which can be mitigated by iron chelation. Together, our work reveals cytoplasmic redox active iron as a novel rheostat in adult stem cells; it demonstrates a role for the intracellular labile iron pool in coordinating a cascade of molecular events which reinforces HSC identity during cell division and to drive stem cell ageing when perturbed. As loss of iron homeostasis is commonly observed in the elderly, we anticipate these findings to trigger further studies into understanding and therapeutic mitigation of labile iron pool-dependent stem cell dysfunction in a wide range of degenerative and malignant pathologies.

cell biology↗

bioRxiv2021

A dynamic and combinatorial histone code drives malaria parasite asexual and sexual development

A histone code defines system-level crosstalk between histone post-translational modifications (PTMs) to induce specific biological outcomes. Proteome-scale information of co-existing PTM across the entire chromatin landscape of the malaria parasite, Plasmodium falciparum, was lacking. Here, we used advanced quantitative middle-down proteomics to identify combinations of PTMs in both the proliferative, asexual stages and transmissible, sexual gametocyte stages of P. falciparum. We provide an updated, high-resolution compendium of 72 PTMs on H3 and H3.3, of which 30 are novel to the parasite. Co-existing PTMs with unique stage distinction was identified, indicating a dynamic and complex histone code with increased connectivity of novel PTMs seen in gametocytes. Chromatin proteomics of a gametocyte-specific combination, H3R17me2K18acK23ac, identified a SAGA-like effector complex (including the transcription factor AP2-G2) tied to this combination to regulate gene expression in mature gametocytes. Ultimately, this study unveils previously undiscovered histone PTMs and their functional relationship with co-existing partners. These results highlight that investigating chromatin regulation in the parasite using single histone PTM assays might overlook higher order gene regulation for distinct proliferation and differentiation processes.

biochemistry↗

bioRxiv2021

VAL genes regulate vegetative phase change via miR156-dependent and independent mechanisms

How organisms control when to transition between different stages of development is a key question in biology. In plants, epigenetic silencing by Polycomb repressive complex 1 (PRC1) and PRC2 plays a crucial role in promoting developmental transitions, including from juvenile-to-adult phases of vegetative growth. PRC1/2 are known to repress the master regulator of vegetative phase change, miR156, leading to the transition to adult growth, but how this process is regulated temporally is unknown. Here we investigate whether transcription factors in the VIVIPAROUS/ABI3-LIKE (VAL) gene family provide the temporal signal for the epigenetic repression of miR156. Exploiting a novel val1 allele, we found that VAL1 and VAL2 redundantly regulate vegetative phase change by controlling the overall level, rather than temporal dynamics, of miR156 expression. Furthermore, we discovered that VAL1 and VAL2 also act independently of miR156 to control this important developmental transition.

plant biology↗