Bridging Integrator 1 Expression Research Articles

BIN1 is essential for β-adrenergic regulation of cardiac function.

The number and activity of voltage gated L-type CaV1.2 Ca2+ channels in the sarcolemma of cardiomyocytes tunes the magnitude of Ca2+-induced Ca2+ release and subsequent contraction. Previous studies from our group revealed that β-adrenergic receptor stimulation in mouse ventricular myocytes triggers mobilization of CaV1.2 channels from early and recycling endosomal reservoirs to augment sarcolemmal expression and clustering and enhancing excitation-contraction coupling. In the heart, the membrane curvature-mediating protein bridging integrator 1 (BIN1) mediates t-tubule micro-folding, and targeted delivery of CaV1.2 channels to t-tubules via anchored microtubules. BIN1 deficits are associated with heart failure. In neurons, BIN1 is known to play an essential role in tubulation of endosomes to permit cargo exit and recycling. We used myocytes isolated from cardiac-specific BIN1 knockout mice, to test the hypothesis that expressional deficits in BIN1 impairs isoproterenol-stimulated endosomal recycling of CaV1.2, reducing the ability to respond to acute stress. Super-resolution Airyscan imaging of immunostained ventricular myocytes revealed that CaV1.2 pools on early and recycling endosomes are mobilized to the sarcolemma to bolster channel expression in response to β-adrenergic stimulation with isoproterenol in WT but not BIN1-/- cells. Moreover, single molecule localization microscopy revealed that isoproterenol-stimulated CaV1.2 upregulation and super-clustering at the t-tubule sarcolemma occurs in wild-type myocytes but is reduced in BIN1+/- and absent in BIN1-/- myocytes. To assess the functional implications on β-adrenergic regulation of CaV1.2 and excitation-contraction coupling, we performed voltage-clamp electrophysiological recordings of whole-cell Ca2+ currents and measured Ca2+transients from paced myocytes finding impaired responses to isoproterenol in BIN1 knockout cells versus wild-type controls. Based on these data, we propose that tight regulation of BIN1 expression is required to support β-adrenergic receptor stimulated CaV1.2 channel trafficking and recycling to increase Ca2+ influx and inotropy in times of acute stress.

Loss of forebrain BIN1 attenuates hippocampal pathology and neuroinflammation in a tauopathy model.

Bridging integrator 1 (BIN1) is the second most prevalent genetic risk factor identified by genome-wide association studies (GWAS) for late-onset Alzheimer's disease. BIN1 encodes an adaptor protein that regulates membrane dynamics in the context of endocytosis and neurotransmitter vesicle release. In vitro evidence suggests that BIN1 can directly bind to tau in the cytosol. In addition, BIN1's function limits extracellular tau seed uptake by endocytosis and subsequent propagation as well as influences tau release through exosomes. However, the in vivo roles of BIN1 in tau pathogenesis and tauopathy-mediated neurodegeneration remain uncharacterized. We generated conditional knockout mice with a selective loss of Bin1 expression in the forebrain excitatory neurons and oligodendrocytes in P301S human tau transgenic background (line PS19). PS19 mice develop age-dependent tau neuropathology and motor deficits and are commonly used to study Alzheimer's disease tau pathophysiology. The severity of motor deficits and neuropathology was compared between experimental and control mice that differ with respect to forebrain BIN1 expression. BIN1's involvement in tau pathology and neuroinflammation was quantified by biochemical methods and immunostaining. Transcriptome changes were profiled by RNA-sequencing analysis to gain molecular insights. The loss of forebrain BIN1 expression in PS19 mice exacerbated tau pathology in the somatosensory cortex, thalamus, spinal cord and sciatic nerve, accelerated disease progression and caused early death. Intriguingly, the loss of BIN1 also mitigated tau neuropathology in select regions, including the hippocampus, entorhinal/piriform cortex, and amygdala, thus attenuating hippocampal synapse loss, neuronal death, neuroinflammation and brain atrophy. At the molecular level, the loss of forebrain BIN1 elicited complex neuronal and non-neuronal transcriptomic changes, including altered neuroinflammatory gene expression, concomitant with an impaired microglial transition towards the disease-associated microglial phenotype. These results provide crucial new information on in vivo BIN1 function in the context of tau pathogenesis. We conclude that forebrain neuronal BIN1 expression promotes hippocampal tau pathogenesis and neuroinflammation. Our findings highlight an exciting region specificity in neuronal BIN1 regulation of tau pathogenesis and reveal cell-autonomous and non-cell-autonomous mechanisms involved in BIN1 modulation of tau neuropathology.

Abstract P495: <Polycystin-1 Regulates Cardiac Bin1 Expression And T-tubule Remodeling>

Background: Transverse tubules (T-tubules) play a key role in cardiac contractility. The expression of Bridging integrator 1 (BIN1), specifically, the cardiac isoforms BIN1+13 and BIN1+ 13+17 promote the formation and ultrastructure of T-tubules and it has been described that the transcriptional factor c-Myc may be a negative regulator of BIN1 expression. Polycystin-1 (PC1) is a mechanosensor in cardiomyocytes, with a crucial role to maintain cardiac function. Our aim was to determine whether PC1 regulates BIN1-induced T-tubule formation by a c-Myc-regulated mechanism. Methods: We used adult C57BL/6 cardiomyocyte-specific knockout mice (PC1 KO). Cardiac function was tested by echocardiography, whereas protein and mRNA content of BIN1, PC1 and c-Myc were measured using western blot and qRT-PCR, respectively. T-tubules were analyzed by transmission electron microscopy (TEM). For statistical analyses, t-test or one-way ANOVA followed by Tukey's test were used. Differences were considered significant at p < 0.05. Results: Survival of PC1 KO mice decreased dramatically after 7 months of age, with clear symptoms of dilated cardiomyopathy and heart failure (decreased fractional shortening and ejection fraction). Ventricular cardiac tissue of PC1 KO mice without (< 7-month-old mice) and with symptoms (7-9-month-old mice) of heart failure (HF) was associated with reduced levels of BIN1 protein content. Total BIN1 mRNA in PC1 KO mice without symptoms did not show differences as compared to controls, but it was significantly decreased in mice with symptoms of HF. Moreover, PC1 KO mice with and without symptoms showed decreased BIN1+13 mRNA levels, whereas BIN1+13+17 mRNA was only increased in mice without symptoms. These changes in BIN1 isoforms were related with T-tubules, which showed increased lumen and decreased intra-luminal density in PC1 KO mice with and without symptoms. Furthermore, c-Myc protein content was increased in cardiac tissue of PC1 KO mice. Conclusion: PC1 may be a regulator of the differential expression of cardiac isoforms of BIN1 in cardiomyocytes by a mechanism that involves negative regulation of c-Myc, related to loss of T-tubule ultrastructure and heart failure development.

The Prognostic Significance of the BIN1 and CCND2 Gene in Adult Patients with Acute Myeloid Leukemia.

To investigate the expression and clinical significance of Bridging INtegrator 1 (BIN1) and cyclin D2 (CCND2) in newly diagnosed cytogenetically heterogenous adult acute myeloid leukemia patients. Methods Real-time quantitative PCR (RQ-PCR) was used to detect the expression of BIN1 and CCND2 genes in 49 newly diagnosed adult patients with AML, and their clinical significance was analyzed. Results BIN1 and CCND2 genes are highly expressed in patients with AML, which suggest their potential as molecular markers. A statistical significant correlation was found between BIN1 expression with bone marrow blasts (P = 0.012), CD34 expression (P = 0.026), NLR (P = 0.019) and Karyotype risk (P = 0.005). There was a statistically significant correlation between CCND2 and age (P = 0.001), NLR (P = 0.046) Karyotype risk (P = 0.033). The expression level of BIN1 was related to early treatment response (CR or not) (P = 0.043). CCND2 expression was significantly correlated with overall survival (P = 0.013), but not with CR (P = 0.731). Conclusion BIN1 and CCND2 genes are highly expressed in patients with AML, suggesting the potential of these two genes as molecular markers. Moreover, high levels of CCND2 are associated with better prognosis, suggesting the possibility of these two genes as prognostic markers.

Prognostic and clinico-pathological significance of BIN1 in breast cancer

Abstract Bridging integrator 1 (BIN1) is a potential tumor suppressor that behaves as silent or misplaced in breast, prostate, skin, lung, and colon cancers. Previous studies noted decreased BIN1 expression in breast cancer (BC). Despite this, BIN1's role in breast cancer is not well understood and remains to be elucidated. Here, we computationally analyzed BIN1 expression and its prognostic role in BC using various databases. Specifically, we found a down-regulated expression profile for the BIN1 gene in different BC subtypes using Oncomine, TCGA, and bcGenExMiner v4.2 web. The expression of BIN1 was found to be associated with various clinico-pathological conditions including estrogen receptor (ER), progesterone receptor (PR) basal-like subtype, and triple-negative breast cancer (TNBC) based on analysis of the bcGenExMiner v4.2 and TCGA databases. In addition to mRNA expression, the correlation between mRNA expression and BIN1 promoter methylation was established using TCGA datasets from UCSC Xena and ULCAN web, and we subsequently examined the relationship of BIN1 promoter methylation with different clinico-pathological conditions in BC patients. Data mining via Kaplan-Meier plotter indicated a positive association between lower BIN1 expression and low BC patient survival and various clinico-pathological conditions in BC patients. Furthermore, we identified a list of genes co-expressed with BIN1 in BC using TCGA data from the ULCAN web. Among the co-expressed genes, glycophorin C (GYPC) had the most predominant positive correlation. This correlation was further confirmed in BC using UCSC Xena and bcGenExMiner v4.2 webs. Overall, this study establishes that attenuated BIN1 expression might induce progression and poor prognosis in BC patients with GYPC association. This could be useful for clinical BC therapy.

SRSF1-dependent alternative splicing attenuates BIN1 expression in non-small cell lung cancer.

Decreased bridging integrator 1 (BIN1) expression has great significance in promoting the progression of malignant tumors. Reduced messenger RNA expression is partly due to aberrant alternative splicing (AS). However, the AS status of BIN1 and its correlation with BIN1 inactivation in non-small cell lung cancer (NSCLC) remains poorly defined. Here we reported that BIN1 inactivation was not related to DNA methylation in NSCLC. Importantly, BIN1 with exon 12A inclusion (BIN1+12A isoform), the most frequent aberrant splicing variant in tumors was also observed in NSCLC, and might be accounted for BIN1 inactivation. Furthermore, we showed that the aberrant splicing of BIN1 was under the control of serine and arginine-rich factor 1 (SRSF1) in NSCLC. In addition, colony formation assay showed that BIN1+12A isoform could abolish the tumor-inhibiting ability of BIN1 in NSCLC cells. Meanwhile, transwell, wound healing and apoptosis experiments demonstrated that theoccurrence of BIN1+12A could abrogate the invasion suppressing activity and proapoptotic property of BIN1 in NSCLC. Significantly, we also found that BIN1+12A isoform neutralized the tumor-suppressing functions of BIN1 via affecting its subcellular localization. Altogether, these data revealed an aberrant splicing phenomenon which abated the expression and tumor-inhibiting activity of BIN1 in NSCLC, and the related mechanisms were associated with SRSF1.

Aberrant accrual of BIN1 near Alzheimer's disease amyloid deposits in transgenic models.

Bridging integrator 1 (BIN1) is the most significant late-onset Alzheimer's disease (AD) susceptibility locus identified via genome-wide association studies. BIN1 is an adaptor protein that regulates membrane dynamics in the context of endocytosis and membrane remodeling. An increase in BIN1 expression and changes in the relative levels of alternatively spliced BIN1 isoforms have been reported in the brains of patients with AD. BIN1 can bind to Tau, and an increase in BIN1 expression correlates with Tau pathology. In contrast, the loss of BIN1 expression in cultured cells elevates Aβ production and Tau propagation by insfluencing endocytosis and recycling. Here, we show that BIN1 accumulates adjacent to amyloid deposits in vivo. We found an increase in insoluble BIN1 and a striking accrual of BIN1 within and near amyloid deposits in the brains of multiple transgenic models of AD. The peri-deposit aberrant BIN1 localization was conspicuously different from the accumulation of APP and BACE1 within dystrophic neurites. Although BIN1 is highly expressed in mature oligodendrocytes, BIN1 association with amyloid deposits occurred in the absence of the accretion of other oligodendrocyte or myelin proteins. Finally, super-resolution microscopy and immunogold electron microscopy analyses highlight the presence of BIN1 in proximity to amyloid fibrils at the edges of amyloid deposits. These results reveal the aberrant accumulation of BIN1 is a feature associated with AD amyloid pathology. Our findings suggest a potential role for BIN1 in extracellular Aβ deposition in vivo that is distinct from its well-characterized function as an adaptor protein in endocytosis and membrane remodeling.

MicroRNA-211 promotes proliferation, migration, and invasion ability of oral squamous cell carcinoma cells via targeting the bridging integrator 1 protein.

Oral squamous cell carcinoma (OSCC), the most common pathological type of oral cancer, is still a frequent malignancy with unsatisfactory prognosis. Accumulating studies have proven some microRNAs (miRNAs) can function as oncogenes in OSCC by targeting tumor suppressors. In this study, we first investigated the expression and role of tumor suppressor bridging integrator‐1 (BIN1) in OSCC tissues and cells. Our results indicated that BIN1 was low expressed in the OSCC tissues and cell lines (SCC6, SCC9, SCC25, HN4, and HN6) along with miR‐211 was highly expressed in OSCC tissues and cell lines, and BIN1 overexpression could evidently inhibit their proliferation, migration, and invasion abilities. Next, we used bioinformation algorithms to predict the potential miRNA targeting BIN1 and chose miR‐211 for further study. miR‐211, a highly expressed miRNA in OSCC cells, could specifically bind with the 3′‐untranslated region (3′‐UTR) of BIN1 to trigger its degradation. Addition of miR‐211 inhibitor could evidently suppress the malignant behaviors of OSCC cells by upregulating BIN1 expression and inhibit the activation of the EGFR/MAPK pathway. Taken together the findings of the study indicated that miR‐211 mediated BIN1 downregulation had crucial significances in OSCC, suggesting the miR‐211 might be a novel potential therapeutic target for the OSCC treatment.

BIN1 reverses PD-L1-mediated immune escape by inactivating the c-MYC and EGFR/MAPK signaling pathways in non-small cell lung cancer.

Non-small cell lung cancer (NSCLC) is one of the most common and malignant carcinoma worldwide, and the incidence and mortality are increasing rapidly. Immunotherapy targeting programmed death 1/programmed death ligand 1 (PD-L1) signaling has shown prominent clinical effects in treating NSCLC; however, a poor understanding of the associated regulating molecular mechanisms of PD-L1 has become one of the biggest obstacles for further improving efficacy. Bridging integrator-1 (BIN1) can regulate numerous cancer-related molecules to exert multiple tumor-suppressing effects by either interacting or not interacting with c-MYC. In the present study, we observed that there exists a negative correlation between the expression of PD-L1 and BIN1 in NSCLC tissues. The expression levels of BIN1 and PD-L1 were significantly related to the tumor, lymph node and metastasis grade (TNM) stage, invasion range and lymph node metastasis. Simultaneously, for NSCLC patients, the expression statuses of BIN1 and PD-L1 might be independent prognostic factors. Furthermore, the expression of tumor-infiltrating lymphocytes was positively associated with BIN1 expression and negatively related to PD-L1 expression in NSCLC tissues. Importantly, we showed that PD-L1 was under the control of BIN1. In addition, the overexpression of BIN1 could inhibit the c-MYC and epithelial growth factor receptor (EGFR)-dependent PD-L1 expression and reverse the suppressive immuno-microenvironment in vivo. Taken together, our findings indicated that BIN1 restoration in NSCLC could reverse PD-L1-mediated immune escape by inactivating the c-MYC and EGFR/mitogen-activated protein kinase pathways.

Inverse regulation of bridging integrator 1 and BCR-ABL1 in chronic myeloid leukemia.

Endocytosis is the major regulator process of tyrosine kinase receptor (RTK) functional activities. Bridging integrator 1 (BIN1) is a key protein involved in RTK intracellular trafficking. Here, we report, by studying 34 patients with chronic myeloid leukemia (CML) at diagnosis, that BIN1 gene is downregulated in CML as compared to healthy controls, suggesting an altered endocytosis of RTKs. Rab interactor 1 (RIN1), an activator of BIN1, displayed a similar behavior. Treatment of 57 patients by tyrosine kinase inhibitors caused, along with BCR-ABL1 inactivation, an increase of BIN1 and RIN1 expression, potentially restoring endocytosis. There was a significant inverse correlation between BIN1-RIN1 and BCR-ABL1 expression. In vitro experiments on both CML and nontumorigenic cell lines treated with Imatinib confirmed these results. In order to provide another proof in favor of BIN1 and RIN1 endocytosis function in CML, we demonstrated that Imatinib induced, in K562 cell line, BIN1-RIN1 upregulation accompanied by a parallel AXL receptor internalization into cytoplasmic compartment. This study shows a novel deregulated mechanism in CML patients, indicating BIN1 and RIN1 as players in the maintenance of the abnormal RTK signaling in this hematological disease.

Amphiphysin 2 (BIN1) in physiology and diseases

Amphiphysin 2, also named bridging integrator-1 (BIN1) or SH3P9, has been recently implicated in rare and common diseases affecting different tissues and physiological functions. BIN1 downregulation is linked to cancer progression and also correlates with ventricular cardiomyopathy and arrhythmia preceding heart failure. Increased BIN1 expression is linked to increased susceptibility for late-onset Alzheimer's disease. In addition, altered splicing may account for the muscle component of myotonic dystrophies, while recessive germinal mutations cause centronuclear myopathy. Despite undoubtedly underlining the relevance of BIN1 in human diseases, the molecular and cellular bases leading to such different diseases are unclear at present. BIN1 is a key regulator of endocytosis and membrane recycling, cytoskeleton regulation, DNA repair, cell cycle progression, and apoptosis. In light of the recent findings on the molecular, cellular, and physiological roles of BIN1, we discuss potential pathological mechanisms and highlight common disease pathways and also tissue-specific regulation. Next challenges will be to validate BIN1 both as a prognostic marker for the related diseases and as a potential therapeutic target.

Bin-1 Expression in Normal Rat Cardiac Myocytes and in Myocytes with Reduced T-Tubule Density Due to Cell Culture or Heart Failure

Bridging Integrator-1 (BIN-1), member of the BAR domain protein superfamily, has been implicated in the formation and maintenance of membrane invaginations in skeletal and cardiac muscle cells. However, it is not clear whether BIN-1 expression and t-tubule density are correlated in all circumstances. The aim of the present study was to investigate the relationship between t-tubule density and BIN-1 expression in (i) fresh atrial and ventricular myocytes (ii) ventricular myocytes cultured for 24-72 hours and (iii) ventricular myocytes from animals with monocrotaline-induced right heart failure (HF). Confocal microscopy was used to assess t-tubule density and BIN1 expression in rat myocytes using di-8-ANEPPS fluorescence or immunofluorescence respectively. In fresh ventricular myocytes, a regular t-tubule network was apparent and BIN-1 exhibited a corresponding striated pattern (n=10). In atrial myocytes t-tubules were typically absent and BIN-1 immunofluorescence was only detectable in the peripheral sarcolemma (n=20). In ventricular myocytes after 24, 48 or 72 hrs cell-culture, t-tubule density was reduced to 61 ± 1.4%, 26 ± 0.6% and 11 ± 0.5% respectively (n=10, 8 and 6, P<0.05), while mean cell area did not change significantly. Correspondingly, BIN1 expression was significantly reduced in ventricular myocytes at 48 (to 17 ± 0.14%, n=7, P<0.05) and 72 hrs (to 16 ± 0.09%; n=5, P<0.05) compared with control. In right ventricular myocytes, from rats with right HF and reduced t-tubule density, BIN1 expression decreased significantly (69 ± 1.3%; n=7, P<0.05) compared with saline-injected rats. These results suggest that where t-tubules are absent (e.g. atrial cells) or in circumstances where t-tubule density is decreased (cell culture, HF) BIN-1 expression is correspondingly reduced. This supports the hypothesis that BIN-1 has an important role in formation and maintenance of t-tubules.

Bridging Integrator 1 (BIN1)-Induced T-Tubule Formation in Cardiomyocytes

The T-tubule system is a branching network of membrane invaginations essential for efficient Ca2+-induced Ca2+ release in myocytes. Significant remodeling of this system has been observed during heart failure (HF), resulting in delayed Ca2+ release at sites lacking T-tubules. Despite its considerable role in HF aetiology, the signaling mechanisms behind T-tubule remodeling remain largely unknown. Bridging integrator 1 (BIN1) has recently drawn considerable interest due to its altered expression during HF and ability to tubulate membranes. In rats examined 6 weeks following myocardial infarction, BIN1 transcription was significantly elevated in animals with compensated or end-stage HF, compared to SHAM operated controls. Although overall T-tubule density was unchanged, confocal imaging of isolated cells stained with di-8-ANEPPS confirmed that the transverse pattern of T-tubules was lost in failing cardiomyocytes, as indicated by reduced peak power in fast Fourier transforms. Interestingly, between-peak power was increased, consistent with the increased fraction of longitudinal tubules visible in failing cardiomyocytes. The role of BIN1 in T-tubule growth was investigated using HL-1 cells, a differentiated murine cardiac cell line lacking both intrinsic BIN1 and T-tubules. When transfected with hBIN1, HL-1 cells developed BIN1-positive invaginations as early as 12 hours following transfection. Whereas BIN1 transcript and protein levels rapidly rose to a peak 24 hours following transfection, BIN1-generated tubules gradually increased in density up until 48 hours after transfection. Additionally, early Ca2+ release at sites containing BIN1-positive tubules indicated that these structures improve Ca2+ release synchrony across the cell. We propose that BIN1 is a crucial regulator of T-tubule development in both healthy and failing cardiomyocytes. In the setting of heart failure, increased BIN1 expression may promote growth of longitudinal tubules that compensate for loss of transverse elements, thereby improving calcium homeostasis in this disease.

Characterization of Bridging Integrator 1 (BIN1) as a Potential Tumor Suppressor and Prognostic Marker in Hepatocellular Carcinoma

It has been shown that bridging integrator 1 (BIN1) can interact with c-myelocytomatosis (c-Myc) oncoprotein in cancer. However, the role of BIN1 in hepatocellular carcinoma (HCC) is not clear. In the present study, we investigated the expression and prognostic role of BIN1 in primary HCC and evaluated the function of BIN1 in hepatocarcinogenesis. Using real-time polymerase chain reaction and Western blot analysis, we found significantly decreased expression of BIN1 in primary HCC tumor tissues (n = 42) compared with adjacent normal tissues and in HCC cell lines. Immunohistochemistry analysis also found decreased BIN1 expression in HCC tumor tissues (n = 117). In clinicopathological analysis, loss of BIN1 expression correlated significantly (P < 0.05) with differentiation scores and tumor size. Importantly, decreased expression of BIN1 in tumors was found to be closely associated with a poor prognosis, and we conclude that BIN1 was an independent prognostic factor in a multivariate analysis. In mechanistic studies, restoring BIN1 expression in BIN1-null HCC cells significantly inhibited cell proliferation and colony formation and induced apoptosis of HCC cells. Furthermore, we found that BIN1 overexpression could significantly suppress the motility and invasion of HCC cells in vitro. Our results indicate that BIN1 may function as a potential tumor suppressor and serve as a novel prognostic marker in HCC patients. The BIN1 molecule might play an important role in tumor growth, cell motility and invasion. Modulation of BIN1 expression may lead to clinical applications of this critical molecule in the control of hepatocellular carcinoma as well as in early and effective diagnosis of this aggressive tumor.

BIN1 is reduced and Cav1.2 trafficking is impaired in human failing cardiomyocytes

Heart failure is a growing epidemic, and a typical aspect of heart failure pathophysiology is altered calcium transients. Normal cardiac calcium transients are initiated by Cav1.2 channels at cardiac T tubules. Bridging integrator 1 (BIN1) is a membrane scaffolding protein that causes Cav1.2 to traffic to T tubules in healthy hearts. The mechanisms of Cav1.2 trafficking in heart failure are not known. To study BIN1 expression and its effect on Cav1.2 trafficking in failing hearts. Intact myocardium and freshly isolated cardiomyocytes from nonfailing and end-stage failing human hearts were used to study BIN1 expression and Cav1.2 localization. To confirm Cav1.2 surface expression dependence on BIN1, patch-clamp recordings were performed of Cav1.2 current in cell lines with and without trafficking-competent BIN1. Also, in adult mouse cardiomyocytes, surface Cav1.2 and calcium transients were studied after small hairpin RNA-mediated knockdown of BIN1. For a functional readout in intact heart, calcium transients and cardiac contractility were analyzed in a zebrafish model with morpholino-mediated knockdown of BIN1. BIN1 expression is significantly decreased in failing cardiomyocytes at both mRNA (30% down) and protein (36% down) levels. Peripheral Cav1.2 is reduced to 42% by imaging, and a biochemical T-tubule fraction of Cav1.2 is reduced to 68%. The total calcium current is reduced to 41% in a cell line expressing a nontrafficking BIN1 mutant. In mouse cardiomyocytes, BIN1 knockdown decreases surface Cav1.2 and impairs calcium transients. In zebrafish hearts, BIN1 knockdown causes a 75% reduction in calcium transients and severe ventricular contractile dysfunction. The data indicate that BIN1 is significantly reduced in human heart failure, and this reduction impairs Cav1.2 trafficking, calcium transients, and contractility.