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BioRxiv : the Preprint Server For... Jun 2024Our sense of hearing is critically dependent on the spiral ganglion neurons (SGNs) that connect the sound receptors in the organ of Corti (OC) to the cochlear nuclei of...
Our sense of hearing is critically dependent on the spiral ganglion neurons (SGNs) that connect the sound receptors in the organ of Corti (OC) to the cochlear nuclei of the hindbrain. Type I SGNs innervate inner hair cells (IHCs) to transmit sound signals, while type II SGNs (SGNIIs) innervate outer hair cells (OHCs) to detect moderate-to-intense sound. During development, SGNII afferents make a characteristic 90-degree turn toward the base of the cochlea and innervate multiple OHCs. It has been shown that the Planar Cell Polarity (PCP) pathway acts non-autonomously to mediate environmental cues in the cochlear epithelium for SGNII afferent turning towards the base. However, the underlying mechanisms are unknown. Here, we present evidence that PCP signaling regulates multiple downstream effectors to influence cell adhesion and the cytoskeleton in cochlear supporting cells (SCs), which serve as intermediate targets of SGNII afferents. We show that the core PCP gene Vangl2 regulates the localization of the small GTPase Rac1 and the cell adhesion molecule Nectin3 at SC-SC junctions through which SGNII afferents travel. Through genetic analysis, we also show that loss of Rac1 or Nectin3 partially phenocopied SGNII peripheral afferent turning defects in mutants, and that Rac1 plays a non-autonomous role in this process in part by regulating PCP protein localization at the SC-SC junctions. Additionally, epistasis analysis indicates that Nectin3 and Rac1 likely act in the same genetic pathway to control SGNII afferent turning. Together, these experiments identify Nectin3 and Rac1 as novel regulators of PCP-directed SGNII axon guidance in the cochlea.
PubMed: 38895287
DOI: 10.1101/2024.06.05.597585 -
BioRxiv : the Preprint Server For... Jun 2024Paclitaxel is a standard of care neoadjuvant therapy for patients with triple negative breast cancer (TNBC); however, it shows limited benefit for locally advanced or...
Paclitaxel is a standard of care neoadjuvant therapy for patients with triple negative breast cancer (TNBC); however, it shows limited benefit for locally advanced or metastatic disease. Here we used a coordinated experimental-computational approach to explore the influence of paclitaxel on the cellular and molecular responses of TNBC cells. We found that escalating doses of paclitaxel resulted in multinucleation, promotion of senescence, and initiation of DNA damage induced apoptosis. Single-cell RNA sequencing (scRNA-seq) of TNBC cells after paclitaxel treatment revealed upregulation of innate immune programs canonically associated with interferon response and downregulation of cell cycle progression programs. Systematic exploration of transcriptional responses to paclitaxel and cancer-associated microenvironmental factors revealed common gene programs induced by paclitaxel, IFNB, and IFNG. Transcription factor (TF) enrichment analysis identified 13 TFs that were both enriched based on activity of downstream targets and also significantly upregulated after paclitaxel treatment. Functional assessment with siRNA knockdown confirmed that the TFs FOSL1, NFE2L2 and ELF3 mediate cellular proliferation and also regulate nuclear structure. We further explored the influence of these TFs on paclitaxel-induced cell cycle behavior via live cell imaging, which revealed altered progression rates through G1, S/G2 and M phases. We found that ELF3 knockdown synergized with paclitaxel treatment to lock cells in a G1 state and prevent cell cycle progression. Analysis of publicly available breast cancer patient data showed that high ELF3 expression was associated with poor prognosis and enrichment programs associated with cell cycle progression. Together these analyses disentangle the diverse aspects of paclitaxel response and identify ELF3 upregulation as a putative biomarker of paclitaxel resistance in TNBC.
PubMed: 38895265
DOI: 10.1101/2024.06.04.596911 -
Diet-microbiome interactions promote enteric nervous system resilience following spinal cord injury.BioRxiv : the Preprint Server For... Jun 2024Spinal cord injury (SCI) results in a plethora of physiological dysfunctions across all body systems, including intestinal dysmotility and atrophy of the enteric nervous...
Spinal cord injury (SCI) results in a plethora of physiological dysfunctions across all body systems, including intestinal dysmotility and atrophy of the enteric nervous system (ENS). Typically, the ENS has capacity to recover from perturbation, so it is unclear why intestinal pathophysiologies persist after traumatic spinal injury. With emerging evidence demonstrating SCI-induced alterations to the gut microbiome composition, we hypothesized that modulation of the gut microbiome could contribute to enteric nervous system recovery after injury. Here, we show that intervention with the dietary fiber, inulin prevents ENS atrophy and limits SCI-induced intestinal dysmotility in mice. However, SCI-associated microbiomes and exposure to specific SCI-sensitive gut microbes are not sufficient to modulate injury-induced intestinal dysmotility. Intervention with microbially-derived short-chain fatty acid (SCFA) metabolites prevents ENS dysfunctions and phenocopies inulin treatment in injured mice, implicating these microbiome metabolites in protection of the ENS. Notably, inulin-mediated resilience is dependent on signaling by the cytokine IL-10, highlighting a critical diet-microbiome-immune axis that promotes ENS resilience following SCI. Overall, we demonstrate that diet and microbially-derived signals distinctly impact recovery of the ENS after traumatic spinal injury. This protective diet-microbiome-immune axis may represent a foundation to uncover etiological mechanisms and future therapeutics for SCI-induced neurogenic bowel.
PubMed: 38895207
DOI: 10.1101/2024.06.06.597793 -
Cell Death & Disease Jun 2024As the second most common malignant tumor in the urinary system, renal cell carcinoma (RCC) is imperative to explore its early diagnostic markers and therapeutic...
As the second most common malignant tumor in the urinary system, renal cell carcinoma (RCC) is imperative to explore its early diagnostic markers and therapeutic targets. Numerous studies have shown that AURKB promotes tumor development by phosphorylating downstream substrates. However, the functional effects and regulatory mechanisms of AURKB on clear cell renal cell carcinoma (ccRCC) progression remain largely unknown. In the current study, we identified AURKB as a novel key gene in ccRCC progression based on bioinformatics analysis. Meanwhile, we observed that AURKB was highly expressed in ccRCC tissue and cell lines and knockdown AURKB in ccRCC cells inhibit cell proliferation and migration in vitro and in vivo. Identified CDC37 as a kinase molecular chaperone for AURKB, which phenocopy AURKB in ccRCC. AURKB/CDC37 complex mediate the stabilization of MYC protein by directly phosphorylating MYC at S67 and S373 to promote ccRCC development. At the same time, we demonstrated that the AURKB/CDC37 complex activates MYC to transcribe CCND1, enhances Rb phosphorylation, and promotes E2F1 release, which in turn activates AURKB transcription and forms a positive feedforward loop in ccRCC. Collectively, our study identified AURKB as a novel marker of ccRCC, revealed a new mechanism by which the AURKB/CDC37 complex promotes ccRCC by directly phosphorylating MYC to enhance its stability, and first proposed AURKB/E2F1-positive feedforward loop, highlighting AURKB may be a promising therapeutic target for ccRCC.
Topics: Humans; Carcinoma, Renal Cell; E2F1 Transcription Factor; Kidney Neoplasms; Cell Cycle Proteins; Cell Line, Tumor; Disease Progression; Phosphorylation; Proto-Oncogene Proteins c-myc; Aurora Kinase B; Cell Proliferation; Animals; Gene Expression Regulation, Neoplastic; Mice, Nude; Mice; Cell Movement; Chaperonins
PubMed: 38890303
DOI: 10.1038/s41419-024-06827-y -
Circulation Jun 2024HIF (hypoxia inducible factor) regulates many aspects of cardiac function. We and others previously showed that chronic HIF activation in the heart in mouse models...
BACKGROUND
HIF (hypoxia inducible factor) regulates many aspects of cardiac function. We and others previously showed that chronic HIF activation in the heart in mouse models phenocopies multiple features of ischemic cardiomyopathy in humans, including mitochondrial loss, lipid accumulation, and systolic cardiac dysfunction. In some settings, HIF also causes the loss of peroxisomes. How, mechanistically, HIF promotes cardiac dysfunction is an open question.
METHODS
We used mice lacking cardiac pVHL (von Hippel-Lindau protein) to investigate how chronic HIF activation causes multiple features of ischemic cardiomyopathy, such as autophagy induction and lipid accumulation. We performed immunoblot assays, RNA sequencing, mitochondrial and peroxisomal autophagy flux measurements, and live cell imaging on hearts and isolated cardiomyocytes. We used CRISPR-Cas9 gene editing in mice to validate a novel mediator of cardiac dysfunction in the setting of chronic HIF activation.
RESULTS
We identify a previously unknown pathway by which cardiac HIF activation promotes the loss of mitochondria and peroxisomes. We found that DEPP1 (decidual protein induced by progesterone 1) is induced under hypoxia in a HIF-dependent manner and localizes inside mitochondria. DEPP1 is both necessary and sufficient for hypoxia-induced autophagy and triglyceride accumulation in cardiomyocytes ex vivo. DEPP1 loss increases cardiomyocyte survival in the setting of chronic HIF activation ex vivo, and whole-body Depp1 loss decreases cardiac dysfunction in hearts with chronic HIF activation caused by loss in vivo.
CONCLUSIONS
Our findings identify DEPP1 as a key component in the cardiac remodeling that occurs with chronic ischemia.
PubMed: 38881449
DOI: 10.1161/CIRCULATIONAHA.123.066628 -
ELife Jun 2024Movement is a key feature of animal systems, yet its embryonic origins are not fully understood. Here, we investigate the genetic basis underlying the embryonic onset of...
Movement is a key feature of animal systems, yet its embryonic origins are not fully understood. Here, we investigate the genetic basis underlying the embryonic onset of movement in focusing on the role played by small non-coding RNAs (microRNAs, miRNAs). To this end, we first develop a quantitative behavioural pipeline capable of tracking embryonic movement in large populations of fly embryos, and using this system, discover that the miRNA plays a role in the emergence of movement. Through the combination of spectral analysis of embryonic motor patterns, cell sorting and RNA in situs, genetic reconstitution tests, and neural optical imaging we define that miR-2b-1 influences the emergence of embryonic movement by exerting actions in the developing nervous system. Furthermore, through the combination of bioinformatics coupled to genetic manipulation of miRNA expression and phenocopy tests we identify a previously uncharacterised (but evolutionarily conserved) chloride channel encoding gene - which we term () - as a genetic target that mechanistically links to the onset of movement. Cell-specific genetic reconstitution of expression in a null miRNA mutant background, followed by behavioural assays and target gene analyses, suggest that affects the emergence of movement through effects in sensory elements of the embryonic circuitry, rather than in the motor domain. Our work thus reports the first miRNA system capable of regulating embryonic movement, suggesting that other miRNAs are likely to play a role in this key developmental process in as well as in other species.
Topics: Animals; MicroRNAs; Drosophila melanogaster; Gene Expression Regulation, Developmental; Movement; Embryo, Nonmammalian; Drosophila; Drosophila Proteins
PubMed: 38869942
DOI: 10.7554/eLife.95209 -
ELife Jun 2024Abnormal lung development can cause congenital pulmonary cysts, the mechanisms of which remain largely unknown. Although the cystic lesions are believed to result...
Abnormal lung development can cause congenital pulmonary cysts, the mechanisms of which remain largely unknown. Although the cystic lesions are believed to result directly from disrupted airway epithelial cell growth, the extent to which developmental defects in lung mesenchymal cells contribute to abnormal airway epithelial cell growth and subsequent cystic lesions has not been thoroughly examined. In the present study using genetic mouse models, we dissected the roles of bone morphogenetic protein (BMP) receptor 1a (Bmpr1a)-mediated BMP signaling in lung mesenchyme during prenatal lung development and discovered that abrogation of mesenchymal disrupted normal lung branching morphogenesis, leading to the formation of prenatal pulmonary cystic lesions. Severe deficiency of airway smooth muscle cells and subepithelial elastin fibers were found in the cystic airways of the mesenchymal knockout lungs. In addition, ectopic mesenchymal expression of BMP ligands and airway epithelial perturbation of the Sox2-Sox9 proximal-distal axis were detected in the mesenchymal knockout lungs. However, deletion of Smad1/5, two major BMP signaling downstream effectors, from the lung mesenchyme did not phenocopy the cystic abnormalities observed in the mesenchymal knockout lungs, suggesting that a Smad-independent mechanism contributes to prenatal pulmonary cystic lesions. These findings reveal for the first time the role of mesenchymal BMP signaling in lung development and a potential pathogenic mechanism underlying congenital pulmonary cysts.
Topics: Animals; Bone Morphogenetic Protein Receptors, Type I; Signal Transduction; Mice; Mice, Knockout; Lung; Mesoderm; Cysts; Bone Morphogenetic Proteins; Lung Diseases; Disease Models, Animal
PubMed: 38856718
DOI: 10.7554/eLife.91876 -
Breast Cancer Research : BCR Jun 2024The aberrant amplification of mammary luminal progenitors is at the origin of basal-like breast cancers associated with BRCA1 mutations. Integrins mediate cell-matrix...
BACKGROUND
The aberrant amplification of mammary luminal progenitors is at the origin of basal-like breast cancers associated with BRCA1 mutations. Integrins mediate cell-matrix adhesion and transmit mechanical and chemical signals that drive epithelial stem cell functions and regulate tumor progression, metastatic reactivation, and resistance to targeted therapies. Consistently, we have recently shown that laminin-binding integrins are essential for the expansion and differentiation of mammary luminal progenitors in physiological conditions. As over-expression of the laminin-binding α6 integrin (Itgα6) is associated with poor prognosis and reduced survival in breast cancer, we here investigate the role of Itgα6 in mammary tumorigenesis.
METHODS
We used Blg-Cre; Brca1; Trp53 mice, a model that phenocopies human basal-like breast cancer with BRCA1 mutations. We generated mutant mice proficient or deficient in Itgα6 expression and followed tumor formation. Mammary tumors and pretumoral tissues were characterized by immunohistochemistry, flow cytometry, RT-qPCR, Western blotting and organoid cultures. Clonogenicity of luminal progenitors from preneoplastic glands was studied in 3D Matrigel cultures.
RESULTS
We show that Itga6 deletion favors activation of p16 cell cycle inhibitor in the preneoplastic tissue. Subsequently, the amplification of luminal progenitors, the cell of origin of Brca1-deficient tumors, is restrained in Itgα6-deficient gland. In addition, the partial EMT program operating in Brca1/p53-deficient epithelium is attenuated in the absence of Itgα6. As a consequence of these events, mammary tumor formation is delayed in Itgα6-deficient mice. After tumor formation, the lack of Itgα6 does not affect tumor growth but rather alters their differentiation, resulting in reduced expression of basal cell markers.
CONCLUSIONS
Our data indicate that Itgα6 has a pro-tumorigenic role in Blg-Cre; Brca1; Trp53 mice developing basal-like mammary tumors. In particular, we reveal that Itgα6 is required for the luminal progenitor expansion and the aberrant partial EMT program that precedes the formation of BRCA1 deficient tumors.
Topics: Animals; Integrin alpha6; Female; Tumor Suppressor Protein p53; Mice; BRCA1 Protein; Breast Neoplasms; Humans; Neoplastic Stem Cells; Cell Proliferation; Stem Cells; Gene Deletion; Cell Transformation, Neoplastic
PubMed: 38835038
DOI: 10.1186/s13058-024-01851-4 -
Molecular Medicine (Cambridge, Mass.) Jun 2024Liver kinase B1 (LKB1) is frequently mutated in lung adenocarcinoma, and its loss contributes to tumor progression.
BACKGROUND
Liver kinase B1 (LKB1) is frequently mutated in lung adenocarcinoma, and its loss contributes to tumor progression.
METHODS
To identify LKB1 downstream genes that promote lung adenocarcinoma aggressiveness, we performed bioinformatical analysis using publicly available datasets.
RESULTS
Rab3B was upregulated in LKB1-depleted lung adenocarcinoma cells and suppressed by LKB1 overexpression. CREB protein was enriched at the promoter of Rab3B in lung cancer cells. Silencing of CREB abrogated the upregulation of Rab3B upon LKB1 loss. Immunohistochemistry revealed the elevated expression of Rab3B in lung adenocarcinomas relative to adjacent normal tissues. Upregulation of Rab3B was significantly associated with lymph node metastasis, advanced tumor stage, and reduced overall survival in lung adenocarcinoma patients. Knockdown of Rab3B suppressed and overexpression of Rab3B promoted the proliferation, colony formation, and migration of lung adenocarcinoma cells in vitro. In a mouse xenograft model, Rab3B depletion restrained and Rab3B overexpression augmented the growth of lung adenocarcinoma tumors. Mechanistically, Rab3B interacted with DDX6 and enhanced its protein stability. Ectopic expression of DDX6 significantly promoted the proliferation, colony formation, and migration of lung adenocarcinoma cells. DDX6 knockdown phenocopied the effects of Rab3B depletion on lung adenocarcinoma cells. Additionally, DDX6 overexpression partially rescued the aggressive phenotype of Rab3B-depleted lung adenocarcinoma cells.
CONCLUSION
LKB1 deficiency promotes Rab3B upregulation via a CREB-dependent manner. Rab3B interacts with and stabilizes DDX6 protein to accelerate lung adenocarcinoma progression. The Rab3B-DDX6 axis may be potential therapeutic target for lung adenocarcinoma.
Topics: Animals; Female; Humans; Male; Mice; Adenocarcinoma of Lung; AMP-Activated Protein Kinase Kinases; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cyclic AMP Response Element-Binding Protein; DEAD-box RNA Helicases; Gene Expression Regulation, Neoplastic; Lung Neoplasms; Protein Serine-Threonine Kinases; Protein Stability
PubMed: 38834947
DOI: 10.1186/s10020-024-00848-1 -
Matrix Biology Plus Jun 2024Although the mechanism for activation of latent TGFβ1 and TGFβ3 is understood to involve the binding of the TGFβ propeptide (LAP) to both an integrin and an insoluble...
Although the mechanism for activation of latent TGFβ1 and TGFβ3 is understood to involve the binding of the TGFβ propeptide (LAP) to both an integrin and an insoluble substrate, the activation of latent TGFβ2 has been unclear because the TGFβ2 LAP does not have the classical integrin binding sequence found in the other two TGFβ isoform LAPs. To assess the potential requirement for covalent linkage with a matrix or cell surface protein for the activation of latent TGFβ2, we generated mice in which the TGFβ2 Cys residue predicted to be involved in binding was mutated to Ser (). We reasoned that, if covalent interaction with a second molecule is required for latent TGFβ2 activation, mutant mice should display a null ()-like phenotype. mice closely phenocopy mice with death in utero between E18 and P1 and with congenital heart and kidney defects similar to those described for mice. The mutant latent TGFβ2 is secreted at levels similar to WT, yet TGFβ signaling monitored as nuclear pSmad2 is suppressed. We conclude that, like latent TGFβ1, latent TGFβ2 activation requires binding to an immobilized matrix or plasma membrane molecule.
PubMed: 38831847
DOI: 10.1016/j.mbplus.2024.100149