Cell biology
Abstract
Oncogene expression can cause replication stress (RS), leading to DNA double-strand breaks (DSB) that require repair through pathways such as homologous recombination, non-homologous end-joining, and microhomology-mediated end-joining (MMEJ). Cyclin D1 (encoded by CCND1) is a well-known oncoprotein overexpressed in cancer; however, its role in RS is unknown. Using mantle cell lymphoma (MCL) as a naturally occurring model of cyclin D1 overexpression, we examined its impact on RS and DSB-repair mechanisms. Cyclin D1 overexpression elevated RS, increased DNA damage, especially during mitosis, and caused specific upregulation of MMEJ. Furthermore, cyclin D1 activates the polymerase theta (POLQ) transcription by binding its promoter loci, driving POLΘ-mediated MMEJ that is essential to withstand cyclin D1-induced RS. Moreover, concurrent ATM deficiency further intensifies RS, enhances POLQ expression and heightens reliance on MMEJ mediated DNA damage repair. Consequently, inhibition of POLΘ in cyclin D1-overexpressed settings further exacerbates RS, causing single-strand DNA gap accumulations and chromosomal instability, ultimately leading to apoptosis, an effect amplified in ATM-deficient cells. Targeting MMEJ via POLΘ inhibition is, therefore, an effective strategy in the context of cyclin D1 overexpression and ATM deficiency and may provide a unique therapeutic approach for treating MCL and other malignancies characterized by similar alterations.
Authors
Jithma Abeykoon, Shuhei Asada, Guangli Zhu, Yuna Hirohashi, Lisa Moreau, Divya R. Iyer, Sirisha Mukkavalli, Kalindi Parmar, Gabriella Zambrano, Lige Jiang, Dongni Yi, Michelle Manske, Kimberly Gwin, Rebecca L. King, James R. Cerhan, Xiaosheng Wu, Zhenkun Lou, Geoffrey I. Shapiro, Thomas Witzig, Alan D'Andrea
Abstract
Sustaining the strong rhythmic interactions between cellular adaptations and environmental cues has been posited as essential for preserving the physiological and behavioral alignment of an organism to the proper phase of the daily light/dark (LD) cycle. Here, we demonstrate that mitochondria and synaptic input organization of suprachiasmatic (SCN) vasoactive intestinal peptide–expressing (VIP-expressing) neurons showed circadian rhythmicity. Perturbed mitochondrial dynamics achieved by conditional ablation of the fusogenic protein mitofusin 2 (Mfn2) in VIP neurons caused disrupted circadian oscillation in mitochondria and synapses in SCN VIP neurons, leading to desynchronization of entrainment to the LD cycle in Mfn2-deficient mice that resulted in an advanced phase angle of their locomotor activity onset, alterations in core body temperature, and sleep-wake amount and architecture. Our data provide direct evidence of circadian SCN clock machinery dependence on high-performance, Mfn2-regulated mitochondrial dynamics in VIP neurons for maintaining the coherence in daily biological rhythms of the mammalian organism.
Authors
Milan Stoiljkovic, Jae Eun Song, Hee-kyung Hong, Heiko Endle, Luis Varela, Jonatas Catarino, Xiao-Bing Gao, Zong-Wu Liu, Peter Sotonyi, Sabrina Diano, Jonathan Cedernaes, Joseph T. Bass, Tamas L. Horvath
Abstract
Neuropathic pain is often comorbid with affective disorders. Synaptic plasticity in anterior cingulate cortex (ACC) is assumed to be a crucial interface for pain perception and emotion. Laminin β1 (LAMB1), a key element of extracellular matrix (ECM) in ACC was recently revealed to convey extracellular alterations to intracellular synaptic plasticity and underlie neuropathic pain and aversive emotion. However, it remains elusive what triggers activity-dependent changes of LAMB1 and ECM remodeling after nerve injury. Here, we uncovered a key role of retinoic acid (RA)/RARB signaling in neuropathic pain and associated anxiodepression via regulation of ECM homeostasis. We showed that nerve injury reduced RA level in the serum and ACC in mice and human, which brought about downregulation of its corresponding receptor, RARB. Overexpressing RARB relieved pain hypersensitivity and comorbid anxiodepression, while silencing RARB exacerbated pain sensitivity and induced anxiodepression. Further mechanistic analysis revealed that RARB maintained ECM homeostasis via transcriptional regulation of LAMB1, reversing abnormal synaptic plasticity and eventually improved neuropathic pain and aversive emotion. Taken together with our previous study, we revealed an intracellular-extracellular-intracellular feedforward regulatory network in modulating pain plasticity. Moreover, we identified cingulate RA/RARB signaling as a promising therapeutic target for treatment of neuropathic pain and associated anxiodepression.
Authors
Zhen-Zhen Li, Wan-Neng Liu, Ke-Xin Liu, Zhi-Wei Dou, Rui Zhao, Yun Chen, Meng-Meng Wang, Tao-Zhi Wang, Fei Wang, Wen-Juan Han, Wen-Guang Chu, Xing-Xing Zheng, Rou-Gang Xie, Hua Yuan, Xiao-Fan Jiang, Xiao-Long Sun, Ceng Luo, Shengxi Wu
Abstract
Cytoplasmic TDP43 mislocalization and aggregation are pathological hallmarks of amyotrophic lateral sclerosis (ALS). However, the initial cellular insults that lead to TDP43 mislocalization remain unclear. In this study, we demonstrate that Nemo-like kinase (NLK) — a proline-directed serine/threonine kinase — promotes the mislocalization of TDP43 and other RNA-binding proteins by disrupting nuclear import. NLK levels are selectively elevated in neurons exhibiting TDP43 mislocalization in ALS patient tissues, while genetic reduction of NLK reduces toxicity in human neuron models of ALS. Our findings suggest that NLK is a promising therapeutic target for neurodegenerative diseases.
Authors
Michael E. Bekier II, Emile S. Pinarbasi, Gopinath Krishnan, Jack J. Mesojedec, Madelaine Hurley, Harisankar Harikumar Sheela, Catherine A. Collins, Layla T. Ghaffari, Martina de Majo, Erik M. Ullian, Mark Koontz, Sarah Coleman, Xingli Li, Elizabeth M.H. Tank, Jacob Waksmacki, Fen-Biao Gao, Sami J. Barmada
Abstract
A subgroup (~20-30%) of castration-resistant prostate cancer (CRPC) aberrantly expresses a gastrointestinal (GI) transcriptome governed by two GI-lineage-restricted transcription factors, HNF1A and HNF4G. In this study, we found that expression of GI transcriptome in CRPC correlates with adverse clinical outcomes to androgen receptor signaling inhibitor treatment and shorter overall survival. Bromo- and extra-terminal domain inhibitors (BETi) downregulated HNF1A, HNF4G, and the GI transcriptome in multiple CRPC models, including cell lines, patient-derived organoids, and patient-derived xenografts, while AR and the androgen-dependent transcriptome were largely spared. Accordingly, BETi selectively inhibited growth of GI transcriptome-positive preclinical models of prostate cancer. Mechanistically, BETi inhibited BRD4 binding at enhancers globally, including both AR and HNF4G bound enhancers while gene expression was selectively perturbed. Restoration of HNF4G expression in the presence of BETi rescued target gene expression without rescuing BRD4 binding. This suggests that inhibition of master transcription factors expression underlies the selective transcriptional effects of BETi.
Authors
Shipra Shukla, Dan Li, Woo Hyun Cho, Dana M. Schoeps, Holly M. Nguyen, Jennifer L. Conner, Marjorie L. Roskes, Anisha Tehim, Gabriella Bayshtok, Mohini R. Pachai, Juan Yan, Nicholas A. Teri, Eric Campeau, Sarah Attwell, Patrick Trojer, Irina Ostrovnaya, Anuradha Gopalan, Ekta Khurana, Eva Corey, Ping Chi, Yu Chen
Abstract
Since the lung is a mucosal barrier organ with a unique immunologic environment, mechanisms of immunoregulation in lung cancer may differ from those of other malignancies. Consistent with this notion, we found that CD8+ T cells play a paradoxical role in facilitating, rather than ameliorating, the growth of multiple lung adenocarcinoma models. These include spontaneous, carcinogen-induced, and transplantable tumor cell line models. Specifically, we found that CD8+ T cells promote homing of CD4+Foxp3+ T regulatory cells to the tumor bed by increasing levels of CCR5 chemokines in the tumor microenvironment in an IFN-γ and TNF-α dependent manner. Contrary to their canonical role, these Th1 cytokines contributed to accelerated growth of murine lung adenocarcinomas while suppressing the growth of other malignancies. Surprisingly, lung cancer cells themselves can serve as a dominant source of IFN-γ, and deletion of this cytokine from cancer cells using CRISPR/Cas-9 decreases tumor growth. Importantly for translational applications, a high level of IFN-γ was also found in human lung cancer patients at both the mRNA and protein level. Our data outlines what we deem a novel and previously undefined lung cancer specific immunoregulatory pathway that may be harnessed to tailor immune based therapy specifically for this malignancy.
Authors
Christina Kratzmeier, Mojtaba Taheri, Zhongcheng Mei, Isabelle Lim, May A. Khalil, Brandon Carter-Cooper, Rachel E. Fanaroff, Chin S. Ong, Eric B. Schneider, Stephanie Chang, Erica Leyder, Dongge Li, Irina G. Luzina, Anirban Banerjee, Alexander Sasha Krupnick
Abstract
Up to 10% of patients with severe early-onset obesity carry pathogenic variants in known obesity-related genes, mostly affecting the leptin-melanocortin pathway. Studying children with severe obesity from consanguineous populations provides a unique opportunity to uncover novel molecular mechanisms. Using whole-exome sequencing, followed by a rigorous analytical and filtration strategy, we identified three different homozygous missense variants in SREK1 (encoding Splicing Regulatory glutamic acid and lysine rich protein) in Pakistani children with severe obesity, from three unrelated consanguineous pedigrees. The wild type SREK1 gene of human induced pluripotent stem cell (iPSC)-derived hypothalamic neurons was individually replaced by each of the three variants and the impact of these changes on global gene expression was studied. Neurons expressing the two variants in the SREK1 RNA recognition domain p.P95L and p.T194M, but not the C-terminally located p.E601K, had markedly reduced expression of the small nucleolar RNA clusters SNORD115 and SNORD116, deficiency of which has been implicated in Prader-Willi syndrome (PWS). In addition to hyperphagic obesity the carriers of these two variants had other features of PWS, such as neonatal hypotonia. In conclusion, homozygous variants in SREK1 result in a subtype of severe early onset obesity sharing features with PWS.
Authors
Sadia Saeed, Anna-Maria Siegert, YC Loraine Tung, Roohia Khanam, Qasim M. Janjua, Jaida Manzoor, Mehdi Derhourhi, Bénédicte Toussaint, Brian Y. H. Lam, Sherine Awad, Emmanuel Vaillant, Emmanuel Buse Falay, Souhila Amanzougarene, Hina Ayesha, Waqas Imran Khan, Nosheen Ramzan, Vladimir Saudek, Stephen O'Rahilly, Anthony P. Goldstone, Muhammad Arslan, Amélie Bonnefond, Philippe Froguel, Giles S.H. Yeo
Abstract
The dystrophin-glycoprotein complex (DGC) is composed of peripheral and integral membrane proteins at the muscle cell membrane that link the extracellular matrix with the intracellular cytoskeleton. While it is well-established that genetic mutations that disrupt the structural integrity of DGC result in numerous muscular dystrophies, the three-dimensional structure of the complex has remained elusive. Two recent elegant cryoEM structures of DGC illuminate its molecular architecture and reveal the unique structural placement of sarcospan (SSPN) within the complex. SSPN, a 25-kDa tetraspanin-like protein, anchors beta-dystroglycan to the beta-, gamma- and delta-sarcoglycan trimer, supporting biochemical studies that SSPN is a core element for DGC assembly and stabilization. Here, we advance these studies by revealing that SSPN provides scaffolding in gamma-sarcoglycanopathies enabling substitution of gamma-sarcoglycan by its homolog, zeta-sarcoglycan, leading to the structural integrity of the DGC and prevention of limb-girdle muscular dystrophy R5. Three-dimensional modeling reveals that zeta-sarcoglycan preserves protein-protein interactions with the sarcospan, sarcoglycans, dystroglycan, and dystrophin. The structural integrity of the complex maintains myofiber attachment to the extracellular matrix and protect the cell membrane from contraction-induced damage. These findings demonstrate that sarcospan prevents limb-girdle muscular dystrophy R5 by remodeling of the sarcoglycan complex composition.
Authors
Ekaterina I. Mokhonova, Daniel Helzer, Ravinder Malik, Hafsa Mamsa, Jackson Walker, Mark Maslanka, Tess S. Fleser, Mohammad H. Afsharinia, Shiheng Liu, Johan Holmberg, Z. Hong Zhou, Eric J. Deeds, Kirk C. Hansen, Elizabeth M. McNally, Rachelle H. Crosbie
Abstract
Vasculopathy is a common hallmark of various fibrotic disorders including systemic sclerosis (SSc), yet its underlying etiology and contribution to fibrogenesis remain ill-defined. In SSc the vasculopathy typically precedes the onset of fibrosis and we observed that this phenomenon is recapitulated in the Snail transgenic mouse model of SSc. The vascular anomalies manifest as deformed vessels, endothelial cell dysfunction and vascular leakage. Our investigation into the underlying mechanism of this phenotype revealed that angiopoietin-like protein 2 (ANGPTL2), secreted by the Snail transgenic keratinocytes, is a principal driver of fibrotic vasculopathy. In endothelial cells, ANGPTL2 upregulates pro-fibrotic genes, downregulates various junctional proteins, and prompts the acquisition of mesenchymal characteristics. Inhibiting endothelial cell junctional instability and consequently vascular leakage with a synthetic analog of the microbial metabolite Urolithin A (UAS03) effectively mitigated the vasculopathy and inhibited fibrogenesis. Thus, ANGPTL2 emerges as a promising early biomarker of the disease and inhibiting the vasculopathy inducing effects of this protein with agents such as UAS03 presents an appealing therapeutic avenue to reduce disease severity. These insights hold the potential to revolutionize the approach to the treatment of fibrotic diseases by targeting the vascular defects.
Authors
Dyuti Saha, Ravi Kiran Annadorai, Sujaya Thannimangalath, Neha P. Shroff, Sunny Kataria, Binita Dam, Abhik Dutta, Akshay Hegde, Ankita Hiwale, Venkatesh Ravula, Shagnik Saha, Lekshmi Minikumari Rahulan, Neha Nigam, Neha Singh, Vikas Agarwal, Praveen K. Vemula, Colin Jamora
Abstract
Castration-resistant prostate cancer (CRPC) marks the advanced and lethal stage of prostate cancer (PCa). TRIM28, also known as KAP1, is a transcriptional regulator recently shown to promote CRPC cell proliferation and xenograft tumor growth. Nonetheless, knowledge gaps persist regarding the mechanisms underlying TRIM28 upregulation in CRPC as well as the genomic targets regulated by TRIM28. Here, we report that TRIM28 is a E2F1 target in CRPC. Using an integrated genomic approach, we have demonstrated that TRIM28 forms a positive feedback loop to promote the transcriptional activation and genomic function of E2F1 independent of retinoblastoma (Rb) status. Furthermore, we identified RSK1 as a kinase that directly phosphorylates TRIM28 at S473, and, as such, RSK1 drives the TRIM28/E2F1 feedback loop. Accordingly, pS473-TRIM28 promotes CRPC progression, which is mitigated by RSK inhibition. In summary, our study reveals a critical role of the RSK1–TRIM28–E2F1 axis in CRPC progression, which may be exploited as a vulnerability in treating Rb-deficient CRPC.
Authors
Miyeong Kim, Jinpeng Liu, Yanquan Zhang, Ruixin Wang, Ryan Goettl, Jennifer Grasso, Derek B. Allison, Chi Wang, Tianyan Gao, Xiaoqi Liu, Ka-Wing Fong
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ISSN: 0021-9738 (print), 1558-8238 (online)