Heart failure (HF) remains a major source of morbidity and mortality in the U.S. The multifunctional Ca2+/calmodulin-dependent kinase II (CaMKII) has emerged as a critical regulator of cardiac hypertrophy and failure, although the mechanisms remain unclear. Previous studies have established that the cytoskeletal protein βIV-spectrin coordinates local CaMKII signaling. Here we sought to determine the role of a spectrin/CaMKII complex in maladaptive remodeling in HF. Chronic pressure overload (6 weeks transaortic constriction, TAC) induced a decrease in cardiac function in WT mice but not in animals expressing truncated βIV-spectrin lacking spectrin/CaMKII interaction (qv3J). Underlying observed differences in function was an unexpected differential regulation of STAT3-related genes in qv3J TAC hearts. In vitro experiments demonstrate that βIV-spectrin serves as a target for CaMKII phosphorylation, which regulates its stability. Cardiac-specific βIV-spectrin knockout (βIV-cKO) mice show STAT3 dysregulation, fibrosis and decreased cardiac function at baseline similar to WT TAC. STAT3 inhibition restored normal cardiac structure and function in βIV-cKO and WT TAC hearts. Our studies identify a novel spectrin-based complex essential for regulation of the cardiac response to chronic pressure overload. We anticipate that strategies targeting the new spectrin-based “statosome” will be effective at suppressing maladaptive remodeling in response to chronic stress.
Sathya D. Unudurthi, Drew M. Nassal, Amara Greer-Short, Nehal J. Patel, Taylor Howard, Xianyao Xu, Birce Onal, Tony Satroplus, Deborah Y. Hong, Cemantha M. Lane, Alyssa Dalic, Sara N. Koenig, Adam C. Lehnig, Lisa A. Baer, Hassan Musa, Kristin I. Stanford, Sakima A. Smith, Peter J. Mohler, Thomas J. Hund
The subthalamic nucleus (STN) is an effective therapeutic target for deep brain stimulation (DBS) for Parkinson’s disease (PD) and histamine level is elevated in the basal ganglia in PD patients. However, the endogenous histaminergic modulation on STN neuronal activities and the neuronal mechanism underlying STN-DBS are unknown. Here we report that STN neuronal firing patterns are more crucial than firing rates for motor control. Histamine excited STN neurons, but paradoxically ameliorated parkinsonian motor deficits, which we attributed to regularizing firing patterns of STN neurons via HCN2 channel coupled to H2 receptor. Intriguingly, DBS increased histamine release in the STN and regularized STN neuronal firing patterns under parkinsonian conditions. HCN2 contributed to the DBS-induced regularization of neuronal firing patterns, suppression of excessive beta oscillations, and alleviation of motor deficits in PD. The results reveal an indispensable role for regularizing STN neuronal firing patterns in amelioration of parkinsonian motor dysfunction and a functional compensation for histamine in parkinsonian basal ganglia circuitry. The findings provide insights into mechanisms of STN-DBS as well as potential therapeutic targets and STN-DBS strategies for PD.
Qian-Xing Zhuang, Guang-Ying Li, Bin Li, Chang-Zheng Zhang, Xiao-Yang Zhang, Kang Xi, Hong-Zhao Li, Jian-Jun Wang, Jing-Ning Zhu
Experimental autoimmune encephalomyelitis (EAE) is an inflammatory demyelinating disease of the central nervous system (CNS), induced by the adoptive transfer of myelin-reactive CD4+ T cells into naïve syngeneic mice. It is widely used as a rodent model of multiple sclerosis (MS). EAE lesion development is initiated when transferred CD4+ T cells access the CNS and are reactivated by local antigen presenting cells (APC) bearing endogenous myelin peptide/ MHC Class II complexes. The identity of the CNS resident, lesion-initiating APC is widely debated. Here we demonstrate that classical dendritic cells (cDC) normally reside in the meninges, brain, and spinal cord in the steady state. These cells are unique among candidate CNS APC in their ability to stimulate naïve, as well as effector, myelin-specific T cells to proliferate and produce pro-inflammatory cytokines directly ex vivo. cDC expanded in the meninges and CNS parenchyma in association with disease progression. Selective depletion of cDC led to a decrease in the number of myelin-primed donor T cells in the CNS and reduced the incidence of clinical EAE by half. Based on our findings, we propose that cDC, and the factors that regulate them, be further investigated as potential therapeutic targets in MS.
David A. Giles, Patrick C. Duncker, Nicole M. Wilkinson, Jesse M. Washnock-Schmid, Benjamin M. Segal
ASXL1 is frequently mutated in myeloid malignancies and is known to co-occur with other gene mutations. However, the molecular mechanisms underlying the leukemogenesis associated with ASXL1 and cooperating mutations remain to be elucidated. Here we report that Asxl1 loss cooperated with haploinsufficiency of Nf1, a negative regulator of the RAS signaling pathway, to accelerate the development of myeloid leukemia in mice. Loss of Asxl1 and Nf1 in hematopoietic stem and progenitor cells resulted in a gain-of-function transcriptional activation of multiple pathways critical for leukemogenesis, such as MYC, NRAS, and BRD4. The hyperactive MYC and BRD4 transcription programs were correlated with elevated H3K4 tri-methylation at the promoter regions of genes involving these pathways. Furthermore, pharmacological inhibition of both MAPK pathway and BET bromodomain prevented leukemia initiation and inhibited disease progression in Asxl1Δ/Δ;Nf1Δ/Δ mice. Concomitant mutations of ASXL1 and RAS pathway genes were associated with aggressive progression of myeloid malignancies in patients. This study sheds light on the understanding of the cooperative effect between epigenetic alterations and signaling pathways in accelerating the progression of myeloid malignancies and provides a rational therapeutic strategy for the treatment of myeloid malignancies with ASXL1 and RAS pathway gene mutations.
Peng Zhang, Fuhong He, Jie Bai, Shohei Yamamoto, Shi Chen, Lin Zhang, Mengyao Sheng, Lei Zhang, Ying Guo, Na Man, Hui Yang, Suyun Wang, Tao Cheng, Stephen D. Nimer, Yuan Zhou, Mingjiang Xu, Qian-Fei Wang, Feng-Chun Yang
The ubiquitin-proteasome system (UPS) degrades a protein molecule via two main steps: ubiquitination and proteasomal degradation. Extraproteasomal ubiquitin receptors are thought to couple the two steps but this proposition has not been tested in vivo with vertebrate animals. More importantly, impaired UPS performance plays a major role in cardiac pathogenesis including myocardial ischemia-reperfusion injury (IRI) but the molecular basis of the UPS impairment remains poorly understood. Ubiquilin1 is a bona fide extra-proteasomal ubiquitin receptor. Here we report that cardiomyocyte-restricted knockout of Ubiquilin1 (Ubqln1-CKO) in mice accumulated a surrogate UPS substrate (GFPdgn) and increased myocardial ubiquitinated proteins without altering proteasome activities, and resulted in a late-onset cardiomyopathy and a significantly shortened lifespan. When subject to regional myocardial ischemia-reperfusion, young Ubqln1-CKO mice showed significantly exacerbated cardiac malfunction and enlarged infarct size and, conversely, mice with transgenic Ubqln1 overexpression displayed attenuated IRI. Furthermore, Ubqln1 overexpression facilitated proteasomal degradation of oxidized proteins and the degradation of a UPS surrogate substrate in cultured cardiomyocytes without increasing autophagic flux. These findings demonstrate that Ubiquilin1 is essential to cardiac ubiquitination-proteasome coupling and that an inadequacy in the coupling represents a major pathogenic factor to myocardial IRI, identifying strengthening the coupling as a potential strategy to reduce IRI.
Chengjun Hu, Yihao Tian, Hongxin Xu, Bo Pan, Erin M. Terpstra, Penglong Wu, Hongmin Wang, Faqian Li, Jinbao Liu, Xuejun Wang
Breast cancer (BrCa) is the malignant tumor that most seriously threatens female health; however, the molecular mechanism underlying its progression remains unclear. Here, we found that conditional deletion of HIC1 in the mouse mammary gland might contribute to premalignant transformation in the early stage of tumor formation. Moreover, the chemokine CXCL14 secreted by HIC1-deleted BrCa cells bound to its novel cognate receptor GPR85 on mammary fibroblasts in the microenvironment and was responsible for activating these fibroblasts via the ERK1/2, Akt, and neddylation pathways, whereas the activated fibroblasts promoted BrCa progression via the induction of the epithelial–mesenchymal transition (EMT) by the CCL17/CCR4 axis. Finally, we confirmed that the HIC1-CXCL14-CCL17 loop was associated with the malignant progression of BrCa. Therefore, the crosstalk between HIC1-deleted BrCa cells and mammary fibroblasts might play a critical role in BrCa development. Taken together, exploring the progression of BrCa from the perspective of microenvironment will be beneficial for identifying the potential prognostic marker of breast tumor and providing the more effective treatment strategy.
Yingying Wang, Xiaoling Weng, Luoyang Wang, Mingang Hao, Yue Li, Lidan Hou, Yu Liang, Tianqi Wu, Mengfei Yao, Guowen Lin, Yiwei Jiang, Guohui Fu, Zhaoyuan Hou, Xiangjun Meng, Jinsong Lu, Jianhua Wang
Hearing loss is a significant public health concern, affecting over 250 million people worldwide. Both genetic and environmental etiologies are linked to hearing loss, but in many cases the underlying cellular pathophysiology is not well understood, highlighting the importance of further discovery. We found that inactivation of the gene, Tmtc4 (transmembrane and tetratricopeptide repeat 4), which was broadly expressed in the mouse cochlea, caused acquired hearing loss in mice. Our data showed Tmtc4 enriched in the endoplasmic reticulum, and that it functioned by regulating Ca2+ dynamics and the unfolded protein response (UPR). Given this genetic linkage of the UPR to hearing loss, we demonstrated a direct link between the more common noise-induced hearing loss (NIHL) and the UPR. These experiments suggested a novel approach to treatment. We demonstrated that the small-molecule UPR and stress response modulator ISRIB (Integrated Stress Response Inhibitor), which activates eIF2B, prevented NIHL in a mouse model. Moreover, in an inverse genetic complementation approach, we demonstrated that mice with homozygous inactivation of both Tmtc4 and Chop had less hearing loss than knockout of Tmtc4 alone. This study implicated a novel mechanism for hearing impairment, highlighting a potential treatment approach for a broad range of human hearing-loss disorders.
Jiang Li, Omar Akil, Stephanie L. Rouse, Conor W. McLaughlin, Ian R. Matthews, Lawrence R. Lustig, Dylan K. Chan, Elliott H. Sherr
Despite the success of T cell checkpoint blockade against melanoma, many “cold” tumors such as prostate cancer remain unresponsive. We find that hypoxic zones are prevalent across pre-clinical prostate cancer and resist T cell infiltration even in the context of CTLA-4 and PD-1 blockade. We show that the hypoxia-activated prodrug TH-302 reduces or eliminates hypoxia in these tumors. Combination therapy with this hypoxia-prodrug and checkpoint blockade cooperate to cure more than 80% of TRAMP-C2 prostate tumors. Immunofluorescence imaging shows that TH-302 drives an influx of T cells into hypoxic zones, which are then amplified by checkpoint blockade. Further, combination therapy reduces myeloid-derived suppressor cell density by more than 50%, and causes a persistent defect in the capacity of the tumor to replenish the granulocytic subset. Spontaneous prostate tumors in TRAMP transgenic mice, which are completely resistant to checkpoint blockade, show minimal adenocarcinoma tumor burden at 36 weeks of age and no evidence of neuroendocrine tumors. Survival of Pb-Cre4, Ptenpc−/−Smad4pc−/− mice with highly aggressive prostate adenocarcinoma is also significantly extended by the combination of hypoxia-prodrug and checkpoint blockade. This combination of hypoxia disruption and T cell checkpoint blockade may render some of the most therapeutically resistant cancers sensitive to immunotherapy.
Priyamvada Jayaprakash, Midan Ai, Arthur Liu, Pratha Budhani, Todd Bartkowiak, Jie Sheng, Casey R. Ager, Courtney Nicholas, Ashvin R. Jaiswal, Yanqiu Sun, Krishna Shah, Sadhana Balasubramanyam, Nan Li, Guocan Wang, Jing Ning, Anna Zal, Tomasz Zal, Michael A. Curran
Lysyl-tRNA synthetase (KRS) functions canonically in cytosolic translational processes. However, KRS is highly expressed in colon cancer, and localizes to distinct cellular compartments upon phosphorylations (i.e., the plasma membranes after T52-phosphorylation and the nucleus after S207-phosphorylation), leading to probably alternative non-canonical functions. It is unknown how other subcellular KRSs crosstalk with environmental cues during cancer progression. Here, we demonstrate that the KRS-dependent metastatic behavior of colon cancer spheroids within three-dimensional gels requires communication between cellular molecules and extracellular soluble factors and neighboring cells. Membranous and nuclear KRS were found to participate in invasive cell dissemination of colon cancer spheroids in three dimensional gels. Cancer spheroids secreted GAS6 via a KRS-dependent mechanism and caused the M2 polarization of macrophages, which activated the neighboring cells via secretion of FGF2/GROα/M-CSF to promote cancer dissemination under environmental remodeling via fibroblast-mediated laminins production. Analyses of tissues from clinical colon cancer patients and Krs–/+ animal models for cancer metastasis supported the roles of KRS, GAS6, and M2 macrophages in KRS-dependent positive feedback between tumors and environmental factors. Altogether, KRS in colon cancer cells remodels the microenvironment to promote metastasis, which can thus be therapeutically targeted at these bidirectional KRS-dependent communications of cancer spheroids with environmental cues.
Seo Hee Nam, Doyeun Kim, Doohyung Lee, Hye-Mi Lee, Dae-Geun Song, Jae Woo Jung, Ji Eon Kim, Hye-Jin Kim, Nam Hoon Kwon, Eun-Kyeong Jo, Sunghoon Kim, Jung Weon Lee
SMAD4 is the only common SMAD (co-SMAD) in transforming growth factor (TGF)-β signaling that usually impedes immune cell activation in the tumor microenvironment. However, here we demonstrated that selective deletion of Smad4 in natural killer (NK) cells actually led to dramatically reduced tumor cell rejection and augmented tumor cell metastases, reduced murine cytomegalovirus clearance, as well as impeded NK cell homeostasis and maturation. This was associated with a downregulation of granzyme B (Gzmb), Kit and Prdm1 in Smad4-deficient NK cells. We further unveiled the mechanism by which SMAD4 promoted Gzmb expression. Gzmb was identified as a direct target of a transcriptional complex formed by SMAD4 and JUNB. A JUNB binding site distinct from that for SMAD4 in the proximal Gzmb promoter was required for transcriptional activation by the SMAD4/JUNB complex. In a Tgfbr2 and Smad4 NK cell-specific double conditional knockout model, SMAD4-mediated events were found to be independent of canonical TGF-β signaling. Our study identifies and mechanistically characterizes unusual functions and pathways for SMAD4 in governing innate immune responses to cancer and viral infection, as well as NK cell development.
Youwei Wang, Jianhong Chu, Ping Yi, Wenjuan Dong, Jennifer N. Saultz, Yufeng Wang, Hongwei Wang, Steven D. Scoville, Jianying Zhang, Lai-Chu Wu, Youcai Deng, Xiaoming He, Bethany L. Mundy-Bosse, Aharon G. Freud, Li-Shu Wang, Michael A. Caligiuri, Jianhua Yu
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