Biology subjects

Prosser, B. L.

Publications and source records attributed to Prosser, B. L..

14 recordsLinked to original sources

bioRxiv2026

Cardiac microtubules mediate transverse (t)-tubule growth and homeostasis

Transverse (t)-tubules enable rapid, synchronous Ca release required for efficient cardiac contraction by bringing L-type Ca channels into close apposition with ryanodine receptors. In heart failure with reduced ejection fraction (HFrEF), t-tubule disorganisation and loss occur alongside cardiac microtubule remodelling, contributing to impaired Ca handling and contractile dysfunction. Despite their canonical function in contraction, how t-tubules develop is unknown. Microtubules support delivery of L-type Ca channels to t-tubules via Amphiphysin-II/BIN1, yet whether microtubules directly regulate t-tubule formation and maintenance is unclear. Here, we investigated a role for microtubules in t-tubule development and homeostasis. Neonatal rat ventricular myocytes (NRVMs), which lack endogenous t-tubules, were used as a reductionist model in which BIN1 overexpression induces nascent membrane tubules. Microtubule depolymerisation with nocodazole before BIN1 overexpression impaired BIN1-driven tubule formation, reducing tubule density and length. Dynein inhibition with EHNA produced similar effects, indicating a requirement for microtubule-based motor activity during tubule elongation. Knockdown of the microtubule +TIP tracking protein CLIP-170 also reduced BIN1-driven tubule density, implicating BIN1-CLIP-170-dependent microtubule capture in tubule initiation. Microtubules were also required to maintain existing tubules. In NRVMs with established BIN1-driven tubules, microtubule depolymerisation, microtubule stabilisation or dynein inhibition each reduced tubule density and length. Consistent with this, acute microtubule depolymerisation or stabilisation disrupted native t-tubule networks in isolated adult sheep left atrial myocytes. Together, these findings identify cardiac microtubules as active regulators of t-tubule architecture. We propose that BIN1-dependent tubule formation requires CLIP-170-mediated microtubule plus-end capture and dynein-dependent elongation, while ongoing microtubule dynamics are necessary to preserve mature t-tubule structure.

physiology↗

bioRxiv2026

Active transport of tRNAs facilitates distributed protein synthesis

Large, polarized cells such as cardiomyocytes, skeletal myofibers, and neurons rely on localized protein synthesis to sustain size, remodel and adapt to stress. The subcellular architecture of these cells is also inherently unfavorable for long-range, diffusion-based transport, which may promote their heavy reliance on active transport mechanisms for the localization of RNA and proteins. Transfer RNAs (tRNAs) function as essential regulators of protein synthesis by linking transcription and translation. Since their discovery in the 1950s, tRNA subcellular distribution has been assumed to occur through passive diffusion. Here, we report that there are pools of tRNAs that depend on the microtubule network for distribution in cardiomyocytes, skeletal myofibers and neurons. Employing dual-color live and fixed-cell super-resolution imaging, we demonstrate that active transport of tRNAs involves hitchhiking on the exterior of endolysosomal vesicles (ELV). We establish that leucyl-tRNA synthetase (LeuRS), the tRNA-binding protein that charges leucine to its cognate tRNA and interacts with Rag GTP on the surface of ELVs, is essential for tRNA transport. Disruption of LeuRS-ELV interactions is sufficient to impair long-range, microtubule-dependent tRNA transport, without affecting mRNA or rRNA transport. We also show that preventing tRNA transport is sufficient to impair translation at sites distal from the nucleus as well as globally impair protein synthesis, ultimately reducing cell size. These findings redefine tRNAs as actively trafficked cargo and establish their directed transport as a fundamental layer of translation regulation required for myocyte homeostasis. Highlights1. Pools of tRNAs require an intact microtubule network for distribution in large, complex cells. 2. tRNAs undergo active transport by hitchhiking on endolysosomal vesicles trafficked by microtubule motors. 3. The tRNA synthetase LeuRS functions as a tRNA-specific adaptor linking tRNAs to endolysosomal vesicles, and disrupting this interaction selectively impairs tRNA, but not mRNA or rRNA transport. 4. Loss of tRNA transport reduces distal translation, global protein synthesis, and cell size.

cell biology↗

bioRxiv2025

CRISPR-mediated transcriptional activation as a mutation-independent therapeutic strategy for SYNGAP1-related intellectual disability

Synaptic Ras GTPase-activating protein (SynGAP) regulates synaptic strength and neuronal signaling, with essential roles in cortical development and synaptic plasticity. Heterozygous loss-of-function variants in SYNGAP1 cause SYNGAP1-related intellectual disability (SRID), a severe neurodevelopmental disorder characterized by epilepsy, developmental delay, and autism. SYNGAP1 mutations often result in haploinsufficiency, providing a strong rationale for gene-targeted therapies. However, no treatment currently addresses the underlying genetic cause of SRID. Here, we developed a CRISPR-mediated transcriptional activation (CRISPRa) approach to upregulate the functional Syngap1 allele in a SRID mouse model. CRISPRa activated Syngap1, normalized SynGAP protein expression and downstream signaling, and rescued working memory deficits. We validated the translational potential of this strategy in human induced pluripotent stem cell (hiPSC)-derived excitatory cortical neurons. CRISPRa rescued SYNGAP1 in two distinct loss-of-function variant lines. Together, these findings demonstrate the feasibility of mutation-independent transcriptional activation as a therapeutic approach for SRID and its broader applicability to haploinsufficiency disorders.

neuroscience↗

bioRxiv2025

Single molecule imaging reveals extracellular signal-regulated kinases dependent signal responsive spatial regulation of translation in cardiomyocytes

BackgroundTranslational control of gene expression is crucial in cardiomyocytes, particularly in response to hypertrophic stimuli. The extracellular signal-regulated kinases (ERK) pathway plays a key role in inducing cardiac hypertrophy and in regulating specific protein translation. However, it is unclear how the specificity is achieved, and the spatiotemporal dynamics of protein translation remain unexplored. MethodsWe employed single-molecule imaging of nascent peptides (SINAPs) reporters to visualize and analyze the translation dynamics in single adult rat ventricular cardiomyocytes and tracked active translation sites at high spatiotemporal resolution. We also examined the effects of adrenergic stimulation and the role of the ERK pathway in translation localization. ResultsOur findings revealed that translation sites are primarily localized near Z-lines in cardiomyocytes, with some sites being highly dynamic and moving during translation. The 3 untranslated regions did not significantly change the localization of translation. Many translation sites co-localized with microtubules, and their movement predominantly occurred along microtubular tracks. Adrenergic stimulation led to a transient shift in translation activity toward the peri-nuclear region, peaking at 12 hours and requiring ERK pathway activity for this localization change. ConclusionsOur high-resolution single-cell study demonstrates that protein translation in cardiomyocytes is dynamic and responsive to hypertrophic stimuli in an ERK-dependent manner. The localized translation mechanism allows cardiomyocytes to rapidly adapt to changing environments by preferentially translating mRNAs in the peri-nuclear region. These findings provide new insights into the spatial regulation of translation in cardiomyocytes and its role in cardiac hypertrophy.

cell biology↗

bioRxiv2025

Dual Translational Control in Cardiomyocytes by Heterogeneous mTORC1 and Hypertrophic ERK Activation

BackgroundCardiac hypertrophy allows post-mitotic cardiomyocytes to meet increased hemodynamic demands but can predispose the heart to adverse clinical outcomes. Despite its central role in cardiac adaptation, the translational control mechanisms that drive cardiac hypertrophy are poorly understood. In this study, we elucidate the relative contributions of the various translational control mechanisms operant during homeostasis and hypertrophic growth. MethodsA combination of immunofluorescence and single myocyte protein synthesis assays were used to dissect the single-cardiomyocyte mechanisms of translational control under basal and hypertrophic conditions in isolated adult rat cardiomyocytes. Translational control mechanism were examined in a mouse model of acute hypertrophic phenylephrine (PE) stimulation prior to overt cardiac growth. ResultsWe observed strikingly heterogeneous activity of mTORC1, the master regulator of translation, across cardiomyocytes both in situ and ex vivo. Heterogeneous mTORC1 activity drove heterogeneous protein synthesis, with translation primarily controlled via canonical mTORC1-dependent 4EBP1 phosphorylation at Thr36/Thr45/Thr69 under baseline conditions. Hypertrophic PE stimulation recruited more cardiomyocytes into a high mTORC1 activity state. PE induced a switch in 4EBP1 phosphorylation by increasing mTORC1-dependent phosphorylation at Thr36/Thr45, but not Thr69. Further, PE induced a novel mTORC1-independent, but MEK-ERK-dependent, pathway driving 4EBP1 phosphorylation at Ser64 in both isolated cardiomyocytes and in vivo. Ribosome biogenesis was also observed within hours upon hypertrophic stimulation, while the mTORC1-S6K-eEF2K-eEF2 pathway was not found to be a major driver of protein translation. ConclusionsProtein synthesis is heterogeneous across cardiomyocytes driven by heterogeneous mTORC1 activity. MEK-ERK signaling directly controls 4EBP1 phosphorylation to augment translation during cardiac hypertrophy and challenges the canonical model of translation initiation. Key pointsO_LImTORC1 activity is low and heterogeneous across cardiomyocytes at baseline. C_LIO_LIHeterogeneous mTORC1 activity drives variable 4EBP1 phosphorylation at Thr36/Thr45/Thr69, resulting in heterogeneous protein translation. C_LIO_LIPhenylephrine stimulation recruits more cardiomyocytes into a high mTORC1 activity state to augment protein synthesis through 4EBP1 phosphorylation at Thr36/Thr45, but not Thr69. C_LIO_LIPhenylephrine stimulation boosts protein translation through mTORC1-independent but ERK-dependent phosphorylation of 4EBP1 at Ser64. C_LIO_LImTORC1-S6K-eEF2K-eEF2 pathway is not a major driver of global protein translation. C_LIO_LIRibosome biogenesis is observed within hours after hypertrophic stimulation. C_LIO_LIThe data demonstrates a dual input model of mTORC1 and ERK dependent phosphorylation of 4EBP1 to regulate protein translation in cardiomyocytes. C_LI

physiology↗

bioRxiv2025

Ultrastructure analysis of mitochondria, lipid droplet and sarcoplasmic reticulum apposition in human heart failure

BackgroundCardiomyocyte structural remodeling is reported as a causal contributor to heart failure (HF) development and progression. Growing evidence highlights the role of organelle apposition in cardiomyocyte function and homeostasis. Disruptions in organelle crosstalk, such as that between the sarcoplasmic reticulum (SR) and mitochondria, are thought to impact numerous cellular processes such as calcium handling and cellular bioenergetics; two processes that are disrupted and implicated in cardiac pathophysiology. While the physical distance between organelles is thought to be essential for homeostatic cardiomyocyte function, whether the interactions and coupling of organelles are altered in human heart failure remains unclear. MethodsHere, we utilized transmission electron microscopy and careful quantification of ultrastructure to characterize the changes in organelle apposition in cardiomyocytes isolated from the hearts of patients diagnosed with various types of HF. Subsequently we employed molecular approaches to examine the expression of proposed organelle tethers. ResultsWe demonstrate that cardiomyocytes isolated from dilated cardiomyopathy, hypertrophic cardiomyopathy and ischemic cardiomyopathy hearts display smaller, more rounded mitochondria, as compared to nonfailing controls. Failing cardiomyocytes also exhibited disrupted SR-mitochondria juxtaposition and changes in the expression of proposed molecular tethers. Further analysis revealed alterations in lipid droplet dynamics including decreased lipid droplet content and less lipid droplets in association with mitochondria in failing cardiomyocytes. ConclusionHere we observed changes in organelle dynamics in cardiomyocytes isolated from heart failure patients diagnosed with differing etiologies. Our results suggest that organelle structure and apposition may be a ubiquitous contributor to human HF progression. RESEARCH PERSPECTIVEWhat is New? O_LIWe provide a detailed analysis of organelle apposition in human heart failure, which has been understudied, and report that that failing human cardiomyocytes display an increase in distance between mitochondria and both the sarcoplasmic reticulum and lipid droplets. C_LIO_LIStructural changes in organelles are correlated with the expression of proposed organelle tethers. C_LIO_LIResource of ultrastructural changes in organelle apposition in human heart failure resulting from various etiologies. C_LI What Questions Should be Addressed next? O_LIThe results from this study provide rationale for causal experimentation to elucidate the contribution of organelle apposition to the progression of heart failure. Future studies examining mechanisms of mitochondrial tethering to the SR or lipid droplet will evaluate specific targets for therapeutic application. C_LI

cell biology↗

bioRxiv2024

Generation of humanized mouse models to support therapeutic development for SYNGAP1 and STXBP1 disorders

Heterozygous variants in SYNGAP1 and STXBP1 cause distinct neurodevelopmental disorders due to haploinsufficiency of essential synaptic proteins. As gene targeted approaches to correct these disorders often target non-conserved genomic regions, thus limiting their clinical translation, we generated humanized mouse models wherein the entire Syngap1 or Stxbp1 loci were replaced with their human counterparts. Stxbp1 humanized mice exhibited impaired viability, while Stxbp1 hybrid mice (Stxbp1Hu/+) were viable and suitable for evaluating target engagement of human-specific therapeutics. Syngap1 humanized mice were viable and successfully crossed with Syngap1 heterozygous mice to produce a Syngap1 humanized-haploinsufficient model (Syngap1Hu/-). Syngap1Hu/- mice displayed haploinsufficient levels of human SYNGAP1, disease-relevant behaviors, and EEG abnormalities including epileptiform activity and generalized slowing. Importantly, parallel analysis in a cohort of patients with SYNGAP1-disorder revealed similar electrophysiological signatures. Finally, we showed that human gene-targeted antisense oligonucleotides modulate human SYNGAP1 expression in Syngap1Hu/- neurons. Together, we describe new models to support pre-clinical therapeutic development for SYNGAP1 and STXBP1 disorders and identify translational biomarkers of SYNGAP1-disorder in mice and humans to benchmark therapeutic testing.

neuroscience↗

bioRxiv2024

Microtubule forces drive nuclear damage in LMNA cardiomyopathy

Nuclear homeostasis requires a balance of forces between the cytoskeleton and nucleus. Mutations in the LMNA gene, which encodes the nuclear envelope proteins lamin A/C, disrupt this balance by weakening the nuclear lamina. This results in nuclear damage in contractile tissues and ultimately muscle disease. Intriguingly, disrupting the LINC complex that connects the cytoskeleton to the nucleus has emerged as a promising strategy to ameliorate LMNA-associated cardiomyopathy. Yet how LINC complex disruption protects the cardiomyocyte nucleus remains unclear. To address this question, we developed an assay to quantify the coupling of cardiomyocyte contraction to nuclear deformation and interrogated its dependence on the nuclear lamina and LINC complex. We found that, surprisingly, the LINC complex was mostly dispensable for transferring contractile strain to the nucleus, and that increased nuclear strain in lamin A/C-deficient cardiomyocytes was not rescued by LINC complex disruption. Instead, LINC complex disruption eliminated the cage of microtubules encircling the nucleus. Disrupting microtubules was sufficient to prevent nuclear damage and rescue cardiac function induced by lamin A/C deficiency. We computationally simulated the stress fields surrounding cardiomyocyte nuclei and show how microtubule forces generate local vulnerabilities that damage lamin A/C-deficient nuclei. Our work pinpoints localized, microtubule-dependent force transmission to the nucleus as a pathological driver and therapeutic target for LMNA- cardiomyopathy. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=131 SRC="FIGDIR/small/579774v5_ufig1.gif" ALT="Figure 1"> View larger version (52K): org.highwire.dtl.DTLVardef@1443daeorg.highwire.dtl.DTLVardef@12d7a8borg.highwire.dtl.DTLVardef@c85900org.highwire.dtl.DTLVardef@14b3c89_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

bioRxiv2023

Reducing microtubule detyrosination improves heart function in HCM mice and human iPSC-engineered heart tissues

RationaleHypertrophic cardiomyopathy (HCM) is the most common cardiac genetic disorder caused by sarcomeric gene variants and associated with left ventricular (LV) hypertrophy and diastolic dysfunction. The role of the microtubule network has recently gained interest with the findings that -tubulin detyrosination (dTyr-tub) is markedly elevated in heart failure. Acute reduction of dTyr-tub by inhibition of the detyrosinase (VASH/SVBP complex) or activation of the tyrosinase (tubulin tyrosine ligase, TTL) markedly improved contractility and reduced stiffness in human failing cardiomyocytes, and thus poses a new perspective for HCM treatment. ObjectiveIn this study, we tested the impact of chronic tubulin tyrosination in a HCM mouse model (Mybpc3-knock-in; KI), in human HCM cardiomyocytes and in SVBP-deficient human engineered heart tissues (EHTs). Methods and ResultsAAV9-mediated TTL transfer was applied in neonatal wild-type (WT) rodents and 3-week-old KI mice and in HCM human induced pluripotent stem cell (hiPSC)-derived cardiomyocytes. We show that i) TTL for 6 weeks dose-dependently reduced dTyr-tub and improved contractility without affecting cytosolic calcium transients in WT cardiomyocytes; ii) TTL for 12 weeks improved diastolic filling, cardiac output and stroke volume and reduced stiffness in KI mice; iii) TTL for 10 days normalized cell hypertrophy in HCM hiPSC-cardiomyocytes; iv) TTL induced a marked transcription and translation of several tubulins and modulated mRNA or protein levels of components of mitochondria, Z-disc, ribosome, intercalated disc, lysosome and cytoskeleton in KI mice; v) SVBP-deficient EHTs exhibited reduced dTyr-tub levels, higher force and faster relaxation than TTL-deficient and WT EHTs. RNA-seq and mass spectrometry analysis revealed distinct enrichment of cardiomyocyte components and pathways in SVBP-KO vs. TTL-KO EHTs. ConclusionThis study provides the first proof-of-concept that chronic activation of tubulin tyrosination in HCM mice and in human EHTs improves heart function and holds promise for targeting the non-sarcomeric cytoskeleton in heart disease.

physiology↗

bioRxiv2022

An unbiased screen identified the Hsp70-BAG3 complex as a regulator of myosin binding protein C3 stability.

Structured AbstractO_ST_ABSObjectiveC_ST_ABSWe aim to identify regulators of myosin binding protein C3 (MyBP-C) protein homeostasis. BackgroundVariants in myosin binding protein C3 (MYBPC3) account for approximately 50% of familial hypertrophic cardiomyopathy (HCM). Most pathogenic variants in MYBPC3 are truncating variants that lead to reduced total levels of MyBP-C protein. Elucidation of the pathways that regulate MyBP-C protein homeostasis could uncover new therapeutic strategies that restore normal protein levels. MethodWe developed a high-throughput screen to identify compounds that can increase or decrease steady-state levels of MyBP-C in an induced pluripotent stem cell cardiomyocyte (iPSC-CM) model derived from a patient with HCM. To normalize results, we also monitored effects on myosin heavy chain (MYH) and focused on those molecules that selectively modulated MyBP-C levels. ResultsScreening a library of 2,426 known biologically active compounds, we identified compounds which either decreased (241/2426, 9.9%) or increased (29/2426, 1.2%) MyBP-C/MYH levels. After a rigorous validation process, including a counter screen for cellular toxicity, two compounds (JG98 and parthenolide) were confirmed as decreasing MyBP-C levels and no compounds were confirmed to increase MyBP-C levels. For further studies, we focused on JG98, which is an allosteric modulator of heat shock protein 70 (Hsp70), inhibiting its interaction with BAG domain co-chaperones. We found that genetic reduction of BAG3 phenocopies treatment with JG98 by reducing MyBP-C protein levels. ConclusionAn unbiased compound screen identified the Hsp70-BAG3 complex as a regulator of MyBP-C stability. Thus, approaches that stimulate this complexs function may be beneficial in the treatment of HCM. Highlights- Hypertrophic cardiomyopathy (HCM) is commonly caused by pathogenic MYBPC3 variants that reduce total wild-type MyBP-C (the protein encoded by MYBPC3). - It is critical to understand the regulators of MyBP-C protein homeostasis to uncover novel therapeutic strategies. - We developed and executed a high-throughput chemical screen in iPSC-CMs to identify compounds which alter steady-state levels of MyBP-C protein, revealing two compounds, JG98 and parthenolide, that significantly reduced MyBP-C levels. - Validation studies suggest that the complex between heat shock protein 70 (Hsp70) and its co-chaperone BAG3 is a dynamic regulator of MyBP-C stability, suggesting that this axis could be a new therapeutic target for HCM.

cell biology↗

bioRxiv2022

Mapping PTBP splicing in human brain identifies targets for therapeutic splice switching including SYNGAP1

Alternative splicing of neuronal genes is controlled in part by the coordinated action of the polypyrimidine tract binding proteins (PTBP1 and PTBP2). While PTBP1 is ubiquitously expressed, PTBP2 is predominantly neuronal, controlling the expression of such targets as DLG4, which encodes PSD95, a protein important in synaptic function whose deficiency causes neurodevelopmental disorders. Here, we fully define the PTBP2 footprint in the human transcriptome using both human brain tissue and neurons derived from human induced pluripotent stem cells (iPSC-neurons). We identify direct PTBP2 binding sites and define PTBP2-dependent alternative splicing events, finding novel targets such as STXBP1 and SYNGAP1, which are synaptic genes also associated with neurodevelopmental disorders. The resultant PTBP2 binding and splicing maps were used to test if PTBP2 binding could be manipulated to increase gene expression in PTBP-targeted genes that cause disease when haploinsufficient. We find that PTBP2 binding to SYNGAP1 mRNA promotes alternative splicing and non-sense mediated decay. Antisense oligonucleotides that disrupt PTBP binding sites on SYNGAP1 redirect splicing and increase gene and protein expression. Collectively, our data provide a comprehensive view of PTBP2-dependent alternative splicing in human neurons and human cerebral cortex, guiding the development of novel therapeutic tools that may benefit a range of neurodevelopmental disorders.

molecular biology↗

bioRxiv2022

Transcriptional, post-transcriptional, and post-translational mechanisms rewrite the tubulin code during cardiac hypertrophy and failure

A proliferated and post-translationally modified microtubule network underlies cellular growth in cardiac hypertrophy and contributes to contractile dysfunction in heart failure. Yet how the heart achieves this modified network is poorly understood. Determining how the "tubulin code" - the permutations of tubulin isoforms and post-translational modifications - is rewritten upon cardiac stress may provide new targets to modulate cardiac remodeling. Further, while tubulin can autoregulate its own expression, it is unknown if autoregulation is operant in the heart or tuned in response to stress. Here we use heart failure patient samples and murine models of cardiac remodeling to interrogate transcriptional, autoregulatory, and post-translational mechanisms that contribute to microtubule network remodeling at different stages of heart disease. We find that autoregulation is operant across tubulin isoforms in the heart and leads to an apparent disconnect in tubulin mRNA and protein levels in heart failure. We also find that within 4 hours of a hypertrophic stimulus and prior to cardiac growth, microtubule detyrosination is rapidly induced to help stabilize the network. This occurs concomitant with rapid transcriptional and autoregulatory activation of specific tubulin isoforms and microtubule motors. Upon continued hypertrophic stimulation, there is an increase in post-translationally modified microtubule tracks and anterograde motors to support cardiac growth, while total tubulin content increases through progressive transcriptional and autoregulatory induction of tubulin isoforms. Our work provides a new model for how the tubulin code is rapidly rewritten to establish a proliferated, stable microtubule network that drives cardiac remodeling, and provides the first evidence of tunable tubulin autoregulation during pathological progression.

molecular biology↗

bioRxiv2021

Desmin intermediate filaments and tubulin detyrosination stabilize growing microtubules in the cardiomyocyte

In heart failure, an increased abundance of post-translationally detyrosinated microtubules stiffens the cardiomyocyte and impedes its contractile function. Detyrosination promotes interactions between microtubules, desmin intermediate filaments and the sarcomere to increase cytoskeletal stiffness, yet the mechanism by which this occurs is unknown. We hypothesized that detyrosination may regulate the growth and shrinkage of dynamic microtubules to facilitate interactions with desmin and the sarcomere. Through a combination of biochemical assays and direct observation of growing microtubule plus-ends in adult cardiomyocytes, we find that desmin is required to stabilize growing microtubules at the sarcomere Z-disk, where desmin also rescue shrinking microtubules from continued depolymerization. Further, reducing detyrosination (tyrosination) promotes frequent depolymerization and inefficient growth of microtubules. This is concomitant with tyrosination promoting the interaction of microtubules with the depolymerizing protein complex of end-binding protein 1 (EB1) and CAP-Gly domain containing linker protein 1 (CLIP1/CLIP170). The futile growth of tyrosinated microtubules reduces their opportunity for stabilizing interactions at the Z-disk, coincident with tyrosination globally reducing microtubule lifetimes and stability. These data provide a model for how intermediate filaments and tubulin detyrosination establish long-lived and physically reinforced microtubules in the cardiomyocyte, and inform on the mechanism of action for therapies that target microtubules for the treatment of cardiac disease.

cell biology↗

bioRxiv2019

Mechanosensing by the lamina protects against nuclear rupture, DNA damage, and cell cycle arrest

Whether cell forces or extracellular matrix (ECM) can impact genome integrity is largely unclear. Here, acute perturbations (~1hr) to actomyosin stress or ECM elasticity cause rapid and reversible changes in lamin-A, DNA damage, and cell cycle. Embryonic hearts, differentiated iPS-cells, and various nonmuscle cell types all show that actomyosin-driven nuclear rupture causes cytoplasmic mis-localization of DNA repair factors and excess DNA damage. Binucleation and micronuclei increase as telomeres shorten, which all favor cell cycle arrest. Deficiencies in lamin-A and repair factors exacerbate these effects, but lamin-A-associated defects are rescued by repair factor overexpression and by contractility modulators in clinical trials. Contractile cells on stiff ECM normally exhibit low phosphorylation and slow degradation of lamin-A by matrix-metalloprotease-2 (MMP2), and inhibition of this lamin-A turnover and also actomyosin contractility is seen to minimize DNA damage. Lamin-A is thus stress-stabilized to mechano-protect the genome.

cell biology↗