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

Heterozygous variants in SYNGAP1 and STXBP1 lead to distinct neurodevelopmental disorders caused by haploinsufficient levels of post-synaptic SYNGAP1 and pre-synaptic STXBP1, which are critical for normal synaptic function. While several gene-targeted therapeutic approaches have proven efficacious in vitro, these often target regions of the human gene that are not conserved in rodents, hindering the pre-clinical development of these compounds and their transition to the clinic. To overcome this limitation, here we generate and characterize Syngap1 and Stxbp1 humanized mouse models in which we replaced the mouse Syngap1 and Stxbp1 gene, respectively, with the human counterpart, including regulatory and non-coding regions. Fully humanized Syngap1 mice present normal viability and can be successfully crossed with currently available Syngap1 haploinsufficiency mouse models to generate Syngap1 humanized haploinsufficient mice. Stxbp1 mice were successfully humanized, yet exhibit impaired viability (particularly males) and reduced STXBP1 protein abundance. Mouse viability could be improved by outcrossing this model to other mouse strains, while Stxbp1 humanized females and hybrid mice can be used to evaluate target engagement of human-specific therapeutics. Overall, these humanized mouse models represent a broadly available tool to further pre-clinical therapeutic development for SYNGAP1 and STXBP1 disorders.

Chronic Activation of Tubulin Tyrosination Improves Heart Function.

RATIONALE: Hypertrophic cardiomyopathy (HCM) is the most common cardiac genetic disorder caused by sarcomeric gene variants and associated with left ventricular hypertrophy and diastolic dysfunction. The role of the microtubule network has recently gained interest with the findings that microtubule detyrosination (dTyr-MT) is markedly elevated in heart failure. Acute reduction of dTyr-MT by inhibition of the detyrosinase (VASH [vasohibin]/SVBP [small VASH-binding protein] complex) or activation of the tyrosinase (TTL [tubulin tyrosine ligase]) markedly improved contractility and reduced stiffness in human failing cardiomyocytes and thus posed a new perspective for HCM treatment. OBJECTIVE: In this study, we tested the impact of chronic tubulin tyrosination in an HCM mouse model (Mybpc3 knock in), in human HCM cardiomyocytes, and in SVBP-deficient human engineered heart tissues (EHTs). METHODS AND RESULTS: Adeno-associated virus serotype 9-mediated TTL transfer was applied in neonatal wild-type rodents, in 3-week-old knock-in mice, and in HCM human induced pluripotent stem cell-derived cardiomyocytes. We show (1) TTL for 6 weeks dose dependently reduced dTyr-MT and improved contractility without affecting cytosolic calcium transients in wild-type cardiomyocytes; (2) TTL for 12 weeks reduced the abundance of dTyr-MT in the myocardium, improved diastolic filling, compliance, cardiac output, and stroke volume in knock-in mice; (3) TTL for 10 days normalized cell area in HCM human induced pluripotent stem cell-derived cardiomyocytes; (4) TTL overexpression activated transcription of tubulins and other cytoskeleton components but did not significantly impact the proteome in knock-in mice; (5) SVBP-deficient EHTs exhibited reduced dTyr-MT levels, higher force, and faster relaxation than TTL-deficient and wild-type EHTs. RNA sequencing and mass spectrometry analysis revealed distinct enrichment of cardiomyocyte components and pathways in SVBP-deficient versus TTL-deficient EHTs. CONCLUSIONS: This 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 nonsarcomeric cytoskeleton in heart disease.

Vasohibin inhibition improves myocardial relaxation in a rat model of heart failure with preserved ejection fraction.

Heart failure with preserved ejection fraction (HFpEF) is a complex syndrome associated with increased myocardial stiffness and cardiac filling abnormalities. Prior studies implicated increased α-tubulin detyrosination, which is catalyzed by the vasohibin enzymes, as a contributor to increased stabilization of the cardiomyocyte microtubule network (MTN) and stiffness in failing human hearts. We explored whether increased MTN detyrosination contributed to impaired diastolic function in the ZSF1 obese rat model of HFpEF and designed a small-molecule vasohibin inhibitor to ablate MTN detyrosination in vivo. Compared with ZSF1 lean and Wistar Kyoto rats, obese rats exhibited increased tubulin detyrosination concomitant with diastolic dysfunction, left atrial enlargement, and cardiac hypertrophy with a preserved left ventricle ejection fraction, consistent with an HFpEF phenotype. Ex vivo myocardial phenotyping assessed cardiomyocyte mechanics and contractility. Vasohibin inhibitor treatment of isolated cardiomyocytes from obese rats resulted in reduced stiffness and faster relaxation. Acute in vivo treatment with vasohibin inhibitor improved diastolic relaxation in ZSF1 obese rats compared with ZSF1 lean and Wistar Kyoto rats. Vasohibin inhibition also improved relaxation in isolated human cardiomyocytes from both failing and nonfailing hearts. Our data suggest the therapeutic potential for vasohibin inhibition to reduce myocardial stiffness and improve relaxation in HFpEF.

Microtubule forces drive nuclear damage in LMNA cardiomyopathy.

Nuclear homeostasis requires a balance of forces between the cytoskeleton and nucleus. Variants in LMNA disrupt this balance by weakening the nuclear lamina, resulting 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 cardiomyopathy. Yet how LINC disruption protects the cardiomyocyte nucleus remains unclear. To address this, we developed an assay to quantify the coupling of cardiomyocyte contraction to nuclear deformation and interrogated its dependence on the lamina and LINC complex. We found that the LINC complex was surprisingly dispensable for transferring the majority of contractile strain into the nucleus, and that increased nuclear strain in Lmna-deficient myocytes was not rescued by LINC disruption. However, LINC disruption eliminated the cage of microtubules encircling the nucleus, and disrupting microtubules was sufficient to prevent nuclear damage induced by LMNA deficiency. Through computational modeling we simulated the mechanical stress fields surrounding cardiomyocyte nuclei and show how microtubule compression exploits local vulnerabilities to damage LMNA-deficient nuclei. Our work pinpoints localized, microtubule-dependent force transmission through the LINC complex as a pathological driver and therapeutic target for LMNA cardiomyopathy.

STXBP1: fast-forward to a brighter future - a patient organization perspective.

Syntaxin-binding protein 1 related disorder (STXBP1-RD) is a rare neurologic disorder associated with global neurodevelopmental delay, intellectual disability, early-onset epilepsy, motor abnormalities, and autism. The underlying pathophysiology stems from a de novo mutation in the STXBP1 gene, which codes for the STXBP1 protein. The STXBP1 protein is involved in synaptic vesicle fusion and neurotransmitter release. Pathogenic variants in the STXBP1 gene generally result in haploinsufficiency, an impairment in neurotransmitter release, and subsequent dysfunction in neuronal communication. The STXBP1 Foundation was founded in 2017 to support families of children with STXBP1-RD and accelerate the development of effective therapies and, ultimately, a cure for the disorder. The Foundation initially supported research aimed at better understanding the complex phenotypic presentation of the disease as well as the development of animal and cellular models usable by the research community to more fully characterize STXBP1 function and disease pathogenicity. In 2023, the Foundation embarked on its STXBP1 Fast Forward Strategic Plan, which includes a prospective natural history study and substantive biomarker work to drive forward the development of new precision therapies for STXBP1-RD.

STXBP1: fast-forward to a brighter future STXBP1-related disorder (STXBP1-RD) is a rare and severe brain condition. It causes delays in development, learning problems, seizures starting at an early age, movement challenges, and sometimes autism. The main problem comes from a new mutation in the STXBP1 gene, which makes a protein needed for brain cells to communicate properly. When this gene doesn't work right, there's not enough of the protein, leading to trouble with brain cell communication. To help families dealing with this disorder and speed up the search for effective therapies, the STXBP1 Foundation started in 2017. At first, they funded studies to understand the disease better and create models for testing treatments. Then, in 2023, they launched the STXBP1 Fast Forward Strategic Plan. This plan includes studying how the disorder progresses naturally and researching markers that could help develop precise treatments for STXBP1-RD.

Accelerating therapeutic development and clinical trial readiness for STXBP1 and SYNGAP1 disorders.

Gene-targeted therapies for genetic neurodevelopmental disorders (NDDs) are becoming a reality. The Center for Epilepsy and Neurodevelopmental Disorders (ENDD) is currently focused on the development of therapeutics for STXBP1 and SYNGAP1 disorders. Here we review the known clinical features of these disorders, highlight the biological role of STXBP1 and SYNGAP1, and discuss our current understanding of pathogenic mechanisms and therapeutic development. Finally, we provide our perspective as scientists and parents of children with NDDs, and comment on the current challenges for both clinical and basic science endeavors.

Integrated Stress Response Potentiates Ponatinib-Induced Cardiotoxicity.

BACKGROUND: Mitochondrial dysfunction is a primary driver of cardiac contractile failure; yet, the cross talk between mitochondrial energetics and signaling regulation remains obscure. Ponatinib, a tyrosine kinase inhibitor used to treat chronic myeloid leukemia, is among the most cardiotoxic tyrosine kinase inhibitors and causes mitochondrial dysfunction. Whether ponatinib-induced mitochondrial dysfunction triggers the integrated stress response (ISR) to induce ponatinib-induced cardiotoxicity remains to be determined. METHODS: Using human induced pluripotent stem cells-derived cardiomyocytes and a recently developed mouse model of ponatinib-induced cardiotoxicity, we performed proteomic analysis, molecular and biochemical assays to investigate the relationship between ponatinib-induced mitochondrial stress and ISR and their role in promoting ponatinib-induced cardiotoxicity. RESULTS: Proteomic analysis revealed that ponatinib activated the ISR in cardiac cells. We identified GCN2 (general control nonderepressible 2) as the eIF2α (eukaryotic translation initiation factor 2α) kinase responsible for relaying mitochondrial stress signals to trigger the primary ISR effector-ATF4 (activating transcription factor 4), upon ponatinib exposure. Mechanistically, ponatinib treatment exerted inhibitory effects on ATP synthase activity and reduced its expression levels resulting in ATP deficits. Perturbed mitochondrial function resulting in ATP deficits then acts as a trigger of GCN2-mediated ISR activation, effects that were negated by nicotinamide mononucleotide, an NAD+ precursor, supplementation. Genetic inhibition of ATP synthase also activated GCN2. Interestingly, we showed that the decreased abundance of ATP also facilitated direct binding of ponatinib to GCN2, unexpectedly causing its activation most likely because of a conformational change in its structure. Importantly, administering an ISR inhibitor protected human induced pluripotent stem cell-derived cardiomyocytes against ponatinib. Ponatinib-treated mice also exhibited reduced cardiac function, effects that were attenuated upon systemic ISRIB administration. Importantly, ISRIB does not affect the antitumor effects of ponatinib in vitro. CONCLUSIONS: Neutralizing ISR hyperactivation could prevent or reverse ponatinib-induced cardiotoxicity. The findings that compromised ATP production potentiates GCN2-mediated ISR activation have broad implications across various cardiac diseases. Our results also highlight an unanticipated role of ponatinib in causing direct activation of a kinase target despite its role as an ATP-competitive kinase inhibitor.

Chronic activation of tubulin tyrosination in HCM mice and human iPSC-engineered heart tissues improves heart function.

Rationale: Hypertrophic 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 -α-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. Objective: In 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 Results: AAV9-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. Conclusion: This 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.

Delineating clinical and developmental outcomes in STXBP1-related disorders.

STXBP1-related disorders are among the most common genetic epilepsies and neurodevelopmental disorders. However, the longitudinal epilepsy course and developmental end points, have not yet been described in detail, which is a critical prerequisite for clinical trial readiness. Here, we assessed 1281 cumulative patient-years of seizure and developmental histories in 162 individuals with STXBP1-related disorders and established a natural history framework. STXBP1-related disorders are characterized by a dynamic pattern of seizures in the first year of life and high variability in neurodevelopmental trajectories in early childhood. Epilepsy onset differed across seizure types, with 90% cumulative onset for infantile spasms by 6 months and focal-onset seizures by 27 months of life. Epilepsy histories diverged between variant subgroups in the first 2 years of life, when individuals with protein-truncating variants and deletions in STXBP1 (n = 39) were more likely to have infantile spasms between 5 and 6 months followed by seizure remission, while individuals with missense variants (n = 30) had an increased risk for focal seizures and ongoing seizures after the first year. Developmental outcomes were mapped using milestone acquisition data in addition to standardized assessments including the Gross Motor Function Measure-66 Item Set and the Grasping and Visual-Motor Integration subsets of the Peabody Developmental Motor Scales. Quantification of end points revealed high variability during the first 5 years of life, with emerging stratification between clinical subgroups. An earlier epilepsy onset was associated with lower developmental abilities, most prominently when assessing gross motor development and expressive communication. We found that individuals with neonatal seizures or early infantile seizures followed by seizure offset by 12 months of life had more predictable seizure trajectories in early to late childhood compared to individuals with more severe seizure presentations, including individuals with refractory epilepsy throughout the first year. Characterization of anti-seizure medication response revealed age-dependent response over time, with phenobarbital, levetiracetam, topiramate and adrenocorticotropic hormone effective in reducing seizures in the first year of life, while clobazam and the ketogenic diet were effective in long-term seizure management. Virtual clinical trials using seizure frequency as the primary outcome resulted in wide range of trial success probabilities across the age span, with the highest probability in early childhood between 1 year and 3.5 years. In summary, we delineated epilepsy and developmental trajectories in STXBP1-related disorders using standardized measures, providing a foundation to interpret future therapeutic strategies and inform rational trial design.

An Unbiased Screen Identified the Hsp70-BAG3 Complex as a Regulator of Myosin-Binding Protein C3.

Variants in the gene myosin-binding protein C3 (MYBPC3) account for approximately 50% of familial hypertrophic cardiomyopathy (HCM), leading to reduced levels of myosin-binding protein C3 (MyBP-C), the protein product made by gene MYBPC3. Elucidation of the pathways that regulate MyBP-C protein homeostasis could uncover new therapeutic strategies. Toward this goal, we screened a library of 2,426 bioactive compounds and identified JG98, an allosteric modulator of heat shock protein 70 that inhibits interaction with Bcl-2-associated athanogene (BAG) domain co-chaperones. JG98 reduces MyBP-C protein levels. Furthermore, genetic reduction of BAG3 phenocopies treatment with JG-98 by reducing MYBP-C protein levels.. Thus, an unbiased compound screen identified the heat shock protein 70-BAG3 complex as a regulator of MyBP-C stability.

Adult human cardiomyocyte mechanics in osteogenesis imperfecta.

Osteogenesis imperfecta (OI) is an extracellular matrix disorder characterized by defects in collagen-1 transport or synthesis, resulting in bone abnormalities. Although reduced collagen in OI hearts has been associated with reduced myocardial stiffness and left ventricular remodeling, its impact on cardiomyocyte (CM) function has not been studied. Here, we explore the tissue-level and CM-level properties of a heart from a deceased organ donor with OI type I. Proteomics and histology confirmed strikingly low expression of collagen 1. Trabecular stretch confirmed low stiffness on the tissue level. However, CMs retained normal viscoelastic properties as revealed by nanoindentation. Interestingly, OI CMs were hypercontractile relative to nonfailing controls after 24 h of culture. In response to 48 h of culture on surfaces with physiological (10 kPa) and pathological (50 kPa) stiffness, OI CMs demonstrated a greater reduction in contractility than nonfailing CMs, suggesting that OI CMs may have an impaired stress response. Levels of detyrosinated α-tubulin, known to be responsive to extracellular stiffness, were reduced in OI CMs. Together these data confirm multiple CM-level adaptations to low stiffness that extend our understanding of OI in the heart and how CMs respond to extracellular stiffness.NEW & NOTEWORTHY In a rare donation of a heart from an individual with osteogenesis imperfecta (OI), we explored cardiomyocyte (CM) adaptations to low stiffness. This represents the first assessment of cardiomyocyte mechanics in OI. The data reveal the hypercontractility of OI CMs with rapid rundown when exposed to acute stiffness challenges, extending our understanding of OI. These data demonstrate that the impact of OI on myocardial mechanics includes cardiomyocyte adaptations beyond known direct effects on the extracellular matrix.

Big tau aggregation disrupts microtubule tyrosination and causes myocardial diastolic dysfunction: from discovery to therapy.

BACKGROUND: Amyloid plaques and neurofibrillary tangles, the molecular lesions that characterize Alzheimer's disease (AD) and other forms of dementia, are emerging as determinants of proteinopathies 'beyond the brain'. This study aims to establish tau's putative pathophysiological mechanistic roles and potential future therapeutic targeting of tau in heart failure (HF). METHODS AND RESULTS: A mouse model of tauopathy and human myocardial and brain tissue from patients with HF, AD, and controls was employed in this study. Tau protein expression was examined together with its distribution, and in vitro tau-related pathophysiological mechanisms were identified using a variety of biochemical, imaging, and functional approaches. A novel tau-targeting immunotherapy was tested to explore tau-targeted therapeutic potential in HF. Tau is expressed in normal and diseased human hearts, in contradistinction to the current oft-cited observation that tau is expressed specifically in the brain. Notably, the main cardiac isoform is high-molecular-weight (HMW) tau (also known as big tau), and hyperphosphorylated tau segregates in aggregates in HF and AD hearts. As previously described for amyloid-beta, the tauopathy phenotype in human myocardium is of diastolic dysfunction. Perturbation in the tubulin code, specifically a loss of tyrosinated microtubules, emerged as a potential mechanism of myocardial tauopathy. Monoclonal anti-tau antibody therapy improved myocardial function and clearance of toxic aggregates in mice, supporting tau as a potential target for novel HF immunotherapy. CONCLUSION: The study presents new mechanistic evidence and potential treatment for the brain-heart tauopathy axis in myocardial and brain degenerative diseases and ageing.

Mapping PTBP2 binding in human brain identifies SYNGAP1 as a target for therapeutic splice switching.

Alternative splicing of neuronal genes is controlled partly by the coordinated action of polypyrimidine tract binding proteins (PTBPs). While PTBP1 is ubiquitously expressed, PTBP2 is predominantly neuronal. Here, we define the PTBP2 footprint in the human transcriptome using brain tissue and human induced pluripotent stem cell-derived neurons (iPSC-neurons). We map PTBP2 binding sites, characterize PTBP2-dependent alternative splicing events, and identify novel PTBP2 targets including SYNGAP1, a synaptic gene whose loss-of-function leads to a complex neurodevelopmental disorder. We find that PTBP2 binding to SYNGAP1 mRNA promotes alternative splicing and nonsense-mediated decay, and that antisense oligonucleotides (ASOs) that disrupt PTBP binding redirect splicing and increase SYNGAP1 mRNA and protein expression. In SYNGAP1 haploinsufficient iPSC-neurons generated from two patients, we show that PTBP2-targeting ASOs partially restore SYNGAP1 expression. Our data comprehensively map PTBP2-dependent alternative splicing in human neurons and cerebral cortex, guiding development of novel therapeutic tools to benefit neurodevelopmental disorders.

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 level of the sarcomere Z-disk, where desmin also rescues shrinking microtubules from continued depolymerization. Further, reducing detyrosination (i.e. tyrosination) below basal levels promotes frequent depolymerization and less efficient 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 dynamic growth and shrinkage of tyrosinated microtubules reduce their opportunity for stabilizing interactions at the Z-disk region, coincident with tyrosination globally reducing microtubule stability. These data provide a model for how intermediate filaments and tubulin detyrosination establish long-lived and physically reinforced microtubules that stiffen the cardiomyocyte and inform both the mechanism of action and therapeutic index for strategies aimed at restoring tyrosination for the treatment of cardiac disease.

Extracellular stiffness induces contractile dysfunction in adult cardiomyocytes via cell-autonomous and microtubule-dependent mechanisms.

The mechanical environment of the myocardium has a potent effect on cardiomyocyte form and function, yet an understanding of the cardiomyocyte responses to extracellular stiffening remains incomplete. We therefore employed a cell culture substrate with tunable stiffness to define the cardiomyocyte responses to clinically relevant stiffness increments in the absence of cell-cell interactions. When cultured on substrates magnetically actuated to mimic the stiffness of diseased myocardium, isolated rat adult cardiomyocytes exhibited a time-dependent reduction of sarcomere shortening, characterized by slowed contraction and relaxation velocity, and alterations of the calcium transient. Cardiomyocytes cultured on stiff substrates developed increases in viscoelasticity and microtubule detyrosination in association with early increases in the α-tubulin detyrosinating enzyme vasohibin-2 (Vash2). We found that knockdown of Vash2 was sufficient to preserve contractile performance as well as calcium transient properties in the presence of extracellular substrate stiffening. Orthogonal prevention of detyrosination by overexpression of tubulin tyrosine ligase (TTL) was also able to preserve contractility and calcium homeostasis. These data demonstrate that a pathologic increment of extracellular stiffness induces early, cell-autonomous remodeling of adult cardiomyocytes that is dependent on detyrosination of α-tubulin.

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 h 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.