Dynamic Subcellular Localization Research Articles

Regulation of Caenorhabditis elegans HLH-30 subcellular localization dynamics: Evidence for a redox-dependent mechanism.

Basic Helix-Loop-Helix (bHLH) transcription factors TFEB/TFE3 and HLH-30 are key regulators of autophagy induction and lysosomal biogenesis in mammals and C. elegans, respectively. While much is known about the regulation of TFEB/TFE3, how HLH-30 subcellular dynamics and transactivation are modulated are yet poorly understood. Thus, elucidating the regulation of C. elegans HLH-30 will provide evolutionary insight into the mechanisms governing the function of bHLH transcription factor family. We report here that HLH-30 is retained in the cytoplasm mainly through its conserved Ser201 residue and that HLH-30 physically interacts with the 14-3-3 protein FTT-2 in this location. The FoxO transcription factor DAF-16 is not required for HLH-30 nuclear translocation upon stress, despite that both proteins partner to form a complex that coordinately regulates several organismal responses. Similar as described for DAF-16, the importin IMB-2 assists HLH-30 nuclear translocation, but constitutive HLH-30 nuclear localization is not sufficient to trigger its distinctive transcriptional response. Furthermore, we identify FTT-2 as the target of diethyl maleate (DEM), a GSH depletor that causes a transient nuclear translocation of HLH-30. Together, our work demonstrates that the regulation of TFEB/TFE3 and HLH-30 family members is evolutionarily conserved and that, in addition to a direct redox regulation through its conserved single cysteine residue, HLH-30 can also be indirectly regulated by a redox-dependent mechanism, probably through FTT-2 oxidation.

Dynamic localization of the chromosomal passenger complex in trypanosomes is controlled by the orphan kinesins KIN-A and KIN-B.

The chromosomal passenger complex (CPC) is an important regulator of cell division, which shows dynamic subcellular localization throughout mitosis, including kinetochores and the spindle midzone. In traditional model eukaryotes such as yeasts and humans, the CPC consists of the catalytic subunit Aurora B kinase, its activator INCENP, and the localization module proteins Borealin and Survivin. Intriguingly, Aurora B and INCENP as well as their localization pattern are conserved in kinetoplastids, an evolutionarily divergent group of eukaryotes that possess unique kinetochore proteins and lack homologs of Borealin or Survivin. It is not understood how the kinetoplastid CPC assembles nor how it is targeted to its subcellular destinations during the cell cycle. Here, we identify two orphan kinesins, KIN-A and KIN-B, as bona fide CPC proteins in Trypanosoma brucei, the kinetoplastid parasite that causes African sleeping sickness. KIN-A and KIN-B form a scaffold for the assembly of the remaining CPC subunits. We show that the C-terminal unstructured tail of KIN-A interacts with the KKT8 complex at kinetochores, while its N-terminal motor domain promotes CPC translocation to spindle microtubules. Thus, the KIN-A:KIN-B complex constitutes a unique 'two-in-one' CPC localization module, which directs the CPC to kinetochores from S phase until metaphase and to the central spindle in anaphase. Our findings highlight the evolutionary diversity of CPC proteins and raise the possibility that kinesins may have served as the original transport vehicles for Aurora kinases in early eukaryotes.

Dynamic localization of the chromosomal passenger complex in trypanosomes is controlled by the orphan kinesins KIN-A and KIN-B

The chromosomal passenger complex (CPC) is an important regulator of cell division, which shows dynamic subcellular localization throughout mitosis, including kinetochores and the spindle midzone. In traditional model eukaryotes such as yeasts and humans, the CPC consists of the catalytic subunit Aurora B kinase, its activator INCENP, and the localization module proteins Borealin and Survivin. Intriguingly, Aurora B and INCENP as well as their localization pattern are conserved in kinetoplastids, an evolutionarily divergent group of eukaryotes that possess unique kinetochore proteins and lack homologs of Borealin or Survivin. It is not understood how the kinetoplastid CPC assembles nor how it is targeted to its subcellular destinations during the cell cycle. Here, we identify two orphan kinesins, KIN-A and KIN-B, as bona fide CPC proteins in Trypanosoma brucei, the kinetoplastid parasite that causes African sleeping sickness. KIN-A and KIN-B form a scaffold for the assembly of the remaining CPC subunits. We show that the C-terminal unstructured tail of KIN-A interacts with the KKT8 complex at kinetochores, while its N-terminal motor domain promotes CPC translocation to spindle microtubules. Thus, the KIN-A:KIN-B complex constitutes a unique ‘two-in-one’ CPC localization module, which directs the CPC to kinetochores from S phase until metaphase and to the central spindle in anaphase. Our findings highlight the evolutionary diversity of CPC proteins and raise the possibility that kinesins may have served as the original transport vehicles for Aurora kinases in early eukaryotes.

Regulated dynamic subcellular GLUT4 localization revealed by proximal proteome mapping in human muscle cells.

Regulation of glucose transport, which is central for control of whole-body metabolism, is determined by the amount of GLUT4 glucose transporter (also known as SLC2A4) in the plasma membrane (PM) of fat and muscle cells. Physiologic signals [such as activated insulin receptor or AMP-activated protein kinase (AMPK)] increase PM GLUT4. Here, we show that the distribution of GLUT4 between the PM and interior of human muscle cells is dynamically maintained, and that AMPK promotes PM redistribution of GLUT4 by regulating exocytosis and endocytosis. Stimulation of exocytosis by AMPK is mediated by Rab10 and the Rab GTPase-activating protein TBC1D4. APEX2 proximity mapping reveals that GLUT4 traverses both PM-proximal and PM-distal compartments in unstimulated muscle cells, further supporting retention of GLUT4 by a constitutive retrieval mechanism. AMPK-stimulated translocation involves GLUT4 redistribution among the same compartments traversed in unstimulated cells, with a significant recruitment of GLUT4 from the Golgi and trans-Golgi network compartments. Our comprehensive proximal protein mapping provides an integrated, high-density, whole-cell accounting of the localization of GLUT4 at a resolution of ∼20 nm that serves as a structural framework for understanding the molecular mechanisms regulating GLUT4 trafficking downstream of different signaling inputs in a physiologically relevant cell type.

First person – Anuttoma Ray

ABSTRACT First Person is a series of interviews with the first authors of a selection of papers published in Journal of Cell Science, helping researchers promote themselves alongside their papers. Anuttoma Ray is first author on ‘ Regulated dynamic subcellular GLUT4 localization revealed by proximal proteome mapping in human muscle cells’, published in JCS. Anuttoma conducted the research described in this article while a Postdoctoral researcher in Dr Timothy McGraw's lab at Department of Biochemistry, Weill Cornell Medicine, Cornell University, New York, USA. She is now an Editor at Cactus Communications, Princeton, NJ, USA, where she leverages her skills and expertise for the scientific community to drive the creation of high-quality, research-based manuscripts for academic publishing.

System-wide analysis of RNA and protein subcellular localization dynamics

Although the subcellular dynamics of RNA and proteins are key determinants of cell homeostasis, their characterization is still challenging. Here we present an integrative framework to simultaneously interrogate the dynamics of the transcriptome and proteome at subcellular resolution by combining two methods: localization of RNA (LoRNA) and a streamlined density-based localization of proteins by isotope tagging (dLOPIT) to map RNA and protein to organelles (nucleus, endoplasmic reticulum and mitochondria) and membraneless compartments (cytosol, nucleolus and cytosolic granules). Interrogating all RNA subcellular locations at once enables system-wide quantification of the proportional distribution of RNA. We obtain a cell-wide overview of localization dynamics for 31,839 transcripts and 5,314 proteins during the unfolded protein response, revealing that endoplasmic reticulum-localized transcripts are more efficiently recruited to cytosolic granules than cytosolic RNAs, and that the translation initiation factor eIF3d is key to sustaining cytoskeletal function. Overall, we provide the most comprehensive overview so far of RNA and protein subcellular localization dynamics.

Endogenous Tagging of Ciliary Genes in Human RPE1 Cells for Live-Cell Imaging.

CRISPR-mediated endogenous tagging of genes provides unique possibilities to explore the function and dynamic subcellular localization of proteins in living cells. Here, we describe experimental strategies for endogenous PCR-tagging of ciliary genes in human RPE1 cells and how image acquisition and analysis of the expressed fluorescently tagged proteins can be utilized to study the dynamic ciliary processes of intraflagellar transport and vesicular trafficking.

Molecular characterization of a novel polerovirus from bitter gourd plants and dynamic subcellular localization of the virus-encoded proteins

Viruses within the Polerovirus genus in the family of Solemoviridae have a single-stranded positive-sense RNA genome of about 5.6–6.2 kb in length. In this study, the bitter gourd leaves showing yellowing and crumple symptoms were collected for small RNAs (sRNAs) sequencing. Analysis of the contigs de novo assembled from sRNA-sequencing data, followed by RT-PCR and cloning, determined the complete viral genome to be 5665 nucleotides. This virus isolate contains conserved ORF3a, ORF3, ORF4, and other typical features of poleroviruses. The P0 protein of this virus isolate shares less than 74.80% amino acid sequence identity with any of the previously characterized poleroviruses, indicating that it should be a novel polerovirus. We name this virus as bitter gourd yellowing crumple virus (BYCV). We further revealed the dynamic subcellular localization and protein accumulation of seven proteins encoded by BYCV in Nicotiana benthamiana plants over time, and these viral proteins displayed specific subcellular localization. Expressing the BYCV P0 protein using a potato virus X vector caused severe symptoms in N. benthamiana plants, while P0 exhibited weak RNA silencing suppression activity. These findings provide an example for investigating the dynamic subcellular localizations of viral proteins and demonstrate that P0 is a critical protein potentially playing an important role in virus infection.

Dynamic subcellular localization of DydA in Dictyostelium cells

DydA plays an important role in chemotaxis, development, and cell growth as an adaptor protein that connects Ras signaling and cytoskeletal rearrangement. DydA is a downstream effector of RasG and is involved in controlling cell polarity and pseudopodia formation during chemoattractant-directed cell migration. To understand the mechanism by which DydA functions on the cell migration, we investigated the dynamic subcellular localization of DydA in response to chemoattractant stimulation and found that DydA rapidly and transiently translocated to the cell cortex through the RA domain and the PRM region in DydA in response to chemoattractant stimulation. The PRM region appears to play a primary role in the translocation of DydA to the cell cortex and in its localization to the actin foci at the bottom of cells. Colocalization experiments of GFP-PRM with RFP-coronin indicated that GFP-PRM preceded GFP-coronin by 2–3 s in response to chemoattractant stimulation. These results suggest that the PRM region plays an indispensable role in relaying upstream regulators, such as RasG, to downstream effectors by mediating the localization of DydA to the cell cortex upon chemoattractant stimulation.

Physiological Condition-Dependent Changes in Ciliary GPCR Localization in the Brain.

Primary cilia are cellular appendages critical for diverse types of Signaling. They are found on most cell types, including cells throughout the CNS. Cilia preferentially localize certain G-protein-coupled receptors (GPCRs) and are critical for mediating the signaling of these receptors. Several of these neuronal GPCRs have recognized roles in feeding behavior and energy homeostasis. Cell and model systems, such as Caenorhabditis elegans and Chlamydomonas, have implicated both dynamic GPCR cilia localization and cilia length and shape changes as key for signaling. It is unclear whether mammalian ciliary GPCRs use similar mechanisms in vivo and under what conditions these processes may occur. Here, we assess two neuronal cilia GPCRs, melanin-concentrating hormone receptor 1 (MCHR1) and neuropeptide-Y receptor 2 (NPY2R), as mammalian model ciliary receptors in the mouse brain. We test the hypothesis that dynamic localization to cilia occurs under physiological conditions associated with these GPCR functions. Both receptors are involved in feeding behaviors, and MCHR1 is also associated with sleep and reward. Cilia were analyzed with a computer-assisted approach allowing for unbiased and high-throughput analysis. We measured cilia frequency, length, and receptor occupancy. We observed changes in ciliary length, receptor occupancy, and cilia frequency under different conditions for one receptor but not another and in specific brain regions. These data suggest that dynamic cilia localization of GPCRs depends on properties of individual receptors and cells where they are expressed. A better understanding of subcellular localization dynamics of ciliary GPCRs could reveal unknown molecular mechanisms regulating behaviors like feeding.

Molecular Dissection of Crz1 and Its Dynamic Subcellular Localization in Cryptococcus neoformans.

Across lower eukaryotes, the transcription factor Crz1 is dephosphorylated by calcineurin, which facilitates Crz1 translocation to the nucleus to regulate gene expression. In the fungal pathogen Cryptococcus neoformans, calcineurin-Crz1 signaling maintains calcium homeostasis, thermotolerance, cell wall integrity, and morphogenesis. How Crz1 distinguishes different stressors and differentially regulates cellular responses is poorly understood. Through monitoring Crz1 subcellular localization over time, we found that Crz1 transiently localizes to granules after exposure to high temperature or calcium. These granules also host the phosphatase calcineurin and Pub1, a ribonucleoprotein stress granule marker, suggesting a role of stress granules in modulating calcineurin-Crz1 signaling. Additionally, we constructed and analyzed an array of Crz1 truncation mutants. We identified the intrinsically disordered regions in Crz1 contribute to proper stress granule localization, nuclear localization, and function. Our results provide the groundwork for further determination of the mechanisms behind the complex regulation of Crz1.

Retention of ERK in the cytoplasm mediates the pluripotency of embryonic stem cells.

The dynamic subcellular localization of ERK1/2 plays an important role in regulating cell fate. Differentiation of mouse embryonic stem cells (mESCs) involves inductive stimulation of ERK1/2, and therefore, inhibitors of the ERK cascade are used to maintain pluripotency. Interestingly, we found that in pluripotent mESCs, ERK1/2 do not translocate to the nucleus either before or after stimulation. This inhibition of nuclear translocation may be dependent on a lack of stimulated ERK1/2 interaction with importin7 rather than a lack of ERK1/2 phosphorylation activating translocation. At late stages of naive-to-primed transition, the action of the translocating machinery is restored, leading to elevation in ERK1/2-importin7 interaction and their nuclear translocation. Importantly, forcing ERK2 into the naive cells' nuclei accelerates their early differentiation, while prevention of the translocation restores stem cells' pluripotency. Theseresults indicate that prevention of nuclear ERK1/2 translocation serves as a safety mechanism for keeping pluripotency of mESCs.

AKTs do not translocate to the nucleus upon stimulation but AKT3 can constitutively signal from the nuclear envelope.

AKT is a central signaling protein kinase that plays a role in the regulation of cellular survival metabolism and cell growth, as well as in pathologies such as diabetes and cancer. Human AKT consists of three isoforms (AKT1-3) that may fulfill different functions. Here, we report that distinct subcellular localization of the isoforms directly influences their activity and function. AKT1 is localized primarily in the cytoplasm, AKT2 in the nucleus, and AKT3 in the nucleus or nuclear envelope. None of the isoforms actively translocates into the nucleus upon stimulation. Interestingly, AKT3 at the nuclear envelope is constitutively phosphorylated, enabling a constant phosphorylation of TSC2 at this location. Knockdown of AKT3 induces moderate attenuation of cell proliferation of breast cancer cells. We suggest that in addition to the stimulation-induced activation of the lysosomal/cytoplasmic AKT1-TSC2 pathway, a subpopulation of TSC2 is constitutively inactivated by AKT3 at the nuclear envelope of transformed cells.

The UPRmt preserves mitochondrial import to extend lifespan.

The mitochondrial unfolded protein response (UPRmt) is dedicated to promoting mitochondrial proteostasis and is linked to extreme longevity. The key regulator of this process is the transcription factor ATFS-1, which, upon UPRmt activation, is excluded from the mitochondria and enters the nucleus to regulate UPRmt genes. However, the repair proteins synthesized as a direct result of UPRmt activation must be transported into damaged mitochondria that had previously excluded ATFS-1 owing to reduced import efficiency. To address this conundrum, we analyzed the role of the import machinery when the UPRmt was induced. Using in vitro and in vivo analysis of mitochondrial proteins, we surprisingly find that mitochondrial import increases when the UPRmt is activated in an ATFS-1-dependent manner, despite reduced mitochondrial membrane potential. The import machinery is upregulated, and an intact import machinery is essential for UPRmt-mediated lifespan extension. ATFS-1 has a weak mitochondrial targeting sequence (MTS), allowing for dynamic subcellular localization during the initial stages of UPRmt activation.

Golgi-Bypass Is a Major Unconventional Route for Translocation to the Plasma Membrane of Non-Apical Membrane Cargoes in Aspergillus nidulans.

Nutrient transporters have been shown to translocate to the plasma membrane (PM) of the filamentous fungus Aspergillus nidulans via an unconventional trafficking route that bypasses the Golgi. This finding strongly suggests the existence of distinct COPII vesicle subpopulations, one following Golgi-dependent conventional secretion and the other directed towards the PM. Here, we address whether Golgi-bypass concerns cargoes other than nutrient transporters and whether Golgi-bypass is related to cargo structure, size, abundance, physiological function, or polar vs. non-polar distribution in the PM. To address these questions, we followed the dynamic subcellular localization of two selected membrane cargoes differing in several of the aforementioned aspects. These are the proton-pump ATPase PmaA and the PalI pH signaling component. Our results show that neosynthesized PmaA and PalI are translocated to the PM via Golgi-bypass, similar to nutrient transporters. In addition, we showed that the COPII-dependent exit of PmaA from the ER requires the alternative COPII coat subunit LstA, rather than Sec24, whereas PalI requires the ER cargo adaptor Erv14. These findings strengthen the evidence of distinct cargo-specific COPII subpopulations and extend the concept of Golgi-independent biogenesis to essential transmembrane proteins, other than nutrient transporters. Overall, our findings point to the idea that Golgi-bypass might not constitute a fungal-specific peculiarity, but rather a novel major and cargo-specific sorting route in eukaryotic cells that has been largely ignored.

TIRAP-mediated activation of p38 MAPK in inflammatory signaling

The role of TIRAP (toll/interleukin-1 receptor (TIR) domain-containing adapter protein) in macrophage inflammatory signalling has been significantly evolved since its discovery in 2001 due to its dynamic nature and subcellular localization to regulate multiple signaling through several protein–protein interactions (PPIs). Structural analysis of these interactions can reveal a better understanding of their conformational dynamics and the nature of their binding. Tyrosine phosphorylation in the TIR domain of TIRAP is very critical for its function. In toll-like receptor (TLR) 4/2 signalling, Bruton's tyrosine kinase (BTK) and Protein kinase C delta (PKCδ) are known to phosphorylate the Y86, Y106, Y159, and Y187 of TIRAP which is crucial for the downstream function of MAPKs (mitogen-activated protein kinases) activation. The objective of this study is to understand the interaction of TIRAP with p38 MAPK through molecular docking and identify the importance of TIRAP tyrosine phosphorylation in p38 MAPK interaction. In this structural study, we performed an in-silico molecular docking using HADDOCK 2.4, pyDockWEB, ClusPro 2.0, and ZDOCK 3.0.2 tools to unravel the interaction between TIRAP and p38 MAPK. Further, manual in-silico phosphorylations of TIRAP tyrosines; Y86, Y106, Y159, and Y187 was created in the Discovery Studio tool to study the conformational changes in protein docking and their binding affinities with p38 MAPK in comparison to non-phosphorylated state. Our molecular docking and 500 ns of molecular dynamic (MD) simulation study demonstrates that the Y86 phosphorylation (pY86) in TIRAP is crucial in promoting the higher binding affinity (∆Gbind) with p38 MAPK. The conformational changes due to the tyrosine phosphorylation mainly at the Y86 site pull the TIRAP closer to the active site in the kinase domain of p38 MAPK and plays a significant role at the interface site which is reversed in its dephosphorylated state. The heatmap of interactions between the TIRAP and p38 MAPK after the MD simulation shows that the TIRAP pY86 structure makes the highest number of stable hydrogen bonds with p38 MAPK residues. Our findings may further be validated in an in-vitro system and would be crucial for targeting the TIRAP and p38 MAPK interaction for therapeutic purposes against the chronic inflammatory response and associated diseases.

CRISPR/Cas9 Gene Editing of HeLa Cells to Tag Proteins with mNeonGreen.

Subcellular localization dynamics of proteins involved in signal transduction processes is crucial in determining the signaling outcome. However, there is very limited information about the localization of endogenous signaling proteins in living cells. For example, biochemical mechanisms underlying the signaling pathway from epidermal growth factor (EGF) receptor (EGFR) to RAS-RAF and ERK1/2/MAPK are well understood, whereas the operational domains of this pathway in the cell remain poorly characterized. Tagging of endogenous components of signaling pathways with fluorescent proteins allows more accurate characterization of their intracellular dynamics at their native expression levels controlled by endogenous regulatory mechanisms, thus avoiding possible tainting effects of overexpression and mistargeting. In this study, we describe methodological approaches to label components of the EGFR-RAS-MAPK pathway, such as Grb2, KRAS, and NRAS, with the fluorescent protein mNeonGreen (mNG) using CRISPR/Cas9 gene-editing, as well as generation of homozygous single-cell clones of the edited target protein.

CERKL, a retinal dystrophy gene, regulates mitochondrial function and dynamics in the mammalian retina

The retina is a highly active metabolic organ that displays a particular vulnerability to genetic and environmental factors causing stress and homeostatic imbalance. Mitochondria constitute a bioenergetic hub that coordinates stress response and cellular homeostasis, therefore structural and functional regulation of the mitochondrial dynamic network is essential for the mammalian retina. CERKL (ceramide kinase like) is a retinal degeneration gene whose mutations cause Retinitis Pigmentosa in humans, a visual disorder characterized by photoreceptors neurodegeneration and progressive vision loss. CERKL produces multiple isoforms with a dynamic subcellular localization. Here we show that a pool of CERKL isoforms localizes at mitochondria in mouse retinal ganglion cells. The depletion of CERKL levels in CerklKD/KO(knockdown/knockout) mouse retinas cause increase of autophagy, mitochondrial fragmentation, alteration of mitochondrial distribution, and dysfunction of mitochondrial-dependent bioenergetics and metabolism. Our results support CERKL as a regulator of autophagy and mitochondrial biology in the mammalian retina.

Rbm24 displays dynamic functions required for myogenic differentiation during muscle regeneration

Skeletal muscle has a remarkable capacity of regeneration after injury, but the regulatory network underlying this repair process remains elusive. RNA-binding proteins play key roles in the post-transcriptional regulation of gene expression and the maintenance of tissue homeostasis and plasticity. Rbm24 regulates myogenic differentiation during early development, but its implication in adult muscle is poorly understood. Here we show that it exerts multiple functions in muscle regeneration. Consistent with its dynamic subcellular localization during embryonic muscle development, Rbm24 also displays cytoplasm to nucleus translocation during C2C12 myoblast differentiation. In adult mice, Rbm24 mRNA is enriched in slow-twitch muscles along with myogenin mRNA. The protein displays nuclear localization in both slow and fast myofibers. Upon injury, Rbm24 is rapidly upregulated in regenerating myofibers and accumulates in the myonucleus of nascent myofibers. Through satellite cell transplantation, we demonstrate that Rbm24 functions sequentially to regulate myogenic differentiation and muscle regeneration. It is required for myogenin expression at early stages of muscle injury and for muscle-specific pre-mRNA alternative splicing at late stages of regeneration. These results identify Rbm24 as a multifaceted regulator of myoblast differentiation. They provide insights into the molecular pathway orchestrating the expression of myogenic factors and muscle functional proteins during regeneration.

Dynamic subcellular localization and transcription activity of the SRF cofactor MKL2 in the striatum are regulated by MAPK.

Dopamine type 1 receptor (D1R) signaling activates protein kinase A (PKA), which then activates mitogen-activated protein kinase (MAPK) through Rap1, in striatal medium spiny neurons (MSNs). MAPK plays a pivotal role in reward-related behavior through the activation of certain transcription factors. How D1R signaling regulates behavior through transcription factors remains largely unknown. CREB-binding protein (CBP) promotes transcription through hundreds of different transcription factors and is also important for reward-related behavior. To identify transcription factors regulated by dopamine signaling in MSNs, we performed a phosphoproteomic analysis using affinity beads coated with CBP. We obtained approximately 40 novel candidate proteins in the striatum of the C57BL/6 mouse brain after cocaine administration. Among them, the megakaryoblastic leukemia-2 (MKL2) protein, a transcriptional coactivator of serum response factor (SRF), was our focus. We found that the interaction between CBP and MKL2 was increased by cocaine administration. Additionally, MKL2, CBP and SRF formed a ternary complex in vivo. The C-terminal domain of MKL2 interacted with CBP-KIX and was phosphorylated by MAPK in COS7 cells. The activation of PKA-MAPK signaling induced the nuclear localization of MKL2 and increased SRF-dependent transcriptional activity in neurons. These results demonstrate that dopamine signaling regulates the interaction of MKL2 with CBP in a phosphorylation-dependent manner and thereby controls SRF-dependent gene expression. Cover Image for this issue:https://doi.org/10.1111/jnc.15067.