Case report: Familial case with autism spectrum and bipolar disorder showing a 20q11.21 microduplication including TM9SF4.

Autism spectrum disorder (ASD) is characterized by multifactorial etiology and high heritability but can be challenging to be diagnosed, especially in cases presenting subthreshold symptoms with no cognitive or language impairment, which may not be identified until adulthood but may occur in family members of subjects with ASD. This study explores the possible correlation between a genomic imbalance and clinical phenotypes in a family case of a proband with ASD, with subjects presenting full-blown or subthreshold ASD and/or mood disorders. Clinical assessments were carried out by means of the Structured Clinical Interview for DSM-5 (SCID-5) disorders, Autism Spectrum Quotient (AQ), Autism Diagnostic Interview-Revised (ADI-R), Autism Diagnostic Observation Schedule Module 2 (ADOS-2), and Adult Autism Subthreshold Spectrum (AdAS Spectrum). The genetic evaluation included array comparative genomic hybridization (array-CGH). The proband was diagnosed with ASD and bipolar disorder type I (BD-I), her twin brothers with ASD and intellectual disability (ID), and her father and sister with BD type II (BD-II) and autism traits. The proband, her father, twin brothers, and older sister showed a microduplication of 350 kb in 20q11.21. In contrast, the proband's mother did not present the microduplication or any mental disorder. This study reports a microduplication that segregates with family members affected by ASD or autistic traits comorbid in some cases with bipolar disorder, and that has never been reported in healthy subjects. Among the genes harbored in this region, the TM9SF4 gene has been recently implicated in risk for ASD.

CircCEMIP promotes anoikis-resistance by enhancing protective autophagy in prostate cancer cells.

BACKGROUND: Circular RNAs (circRNAs) are essential participants in the development and progression of various malignant tumors. Previous studies have shown that cell migration-inducing protein (CEMIP) accelerates prostate cancer (PCa) anoikis resistance (AR) by activating autophagy. This study focused on the effect of circCEMIP on PCa metastasis. METHODS: This study gradually revealed the role of circ_0004585 in PCa anoikis resistance via quantitative real-time PCR (qRT-PCR) analysis, western blotting, pull-down assays, and dual fluorescence reporter assays. RESULTS: Functionally, circ_0004585 promoted PCa cells invasion and metastasis both in vitro and in vivo. Mechanistically, circ_0004585 directly interacted with miR-1248 to upregulate target gene expression. Furthermore, target prediction and dual-luciferase reporter assays identified transmembrane 9 superfamily member 4 (TM9SF4) as a potential miR-1248 target. Pathway analysis revealed that TM9SF4 activated autophagy to promote PCa cells anoikis resistance via mTOR phosphorylation. CONCLUSIONS: These results demonstrated that circ_0004585 played an oncogenic role during PCa invasion and metastasis by targeting the miR-1248/TM9SF4 axis while providing new insight into therapeutic strategy development for metastatic PCa.

Long Noncoding RNA GUSBP11 Knockdown Alleviates Nasopharyngeal Carcinoma via Regulating miR-1226-3p/TM9SF4 Axis.

Purpose: Long noncoding RNAs (lncRNAs) have been confirmed related to the occurrence and progress of multiple cancers, including cervical cancer nasopharyngeal carcinoma (NPC). This study focused on assessing GUSBP11 effects on NPC progression and exploring possible mechanisms. Materials and Methods: RT-qPCR was conducted for assessing GUSBP11 levels within NPC tissues and cells. CCK-8, colony formation, and Transwell were adopted for examining GUSBP11 impacts on NPC cell proliferation and cell metastasis. RT-qPCR analysis and dual-luciferase reporter assay were conducted for judging the expression interrelation of GUSBP11 and its potential target miR-1226-3p. The same methods were carried out for verifying the inhibiting influences of miR-1226-3p upregulation and its potential target TM9SF4. Results: GUSBP11 levels were upregulated within NPC tissues and cells. GUSBP11 downregulation repressed NPC cell proliferation and cell metastasis. In addition, GUSBP11 targeted and negatively regulated miR-1226-3p. Furthermore, miR-1226-3p targeted TM9SF4 and mediated GUSBP11's impacts on TM9SF4 levels. At last, the authors proved the critical role of the GUSBP11/miR-1226-3p/TM9SF4 axis in regulating NPC progression. Conclusion: These findings indicate that downregulation of GUSBP11 alleviates NPC development by regulating the miR-1226-3p/TM9SF4 axis.

TM9SF4 Is a Crucial Regulator of Inflammation and ER Stress in Inflammatory Bowel Disease.

BACKGROUND & AIMS: Inflammatory bowel disease (IBD) is a major intestinal disease. Excessive inflammation and increased endoplasmic reticulum (ER) stress are the key events in the development of IBD. Search of a genome-wide association study database identified a remarkable correlation between a TM9SF4 single-nucleotide polymorphism and IBD. Here, we aimed to resolve its underlying mechanism. METHODS: The role of TM9SF4 was determined with experimental mouse models of IBD. ER stress cascades, barrier functions, and macrophage polarization in colonic tissues and cells were assessed in vivo and in vitro. The expression of TM9SF4 was compared between inflamed regions of ulcerative colitis patients and normal colon samples. RESULTS: In mouse models of IBD, genetic knockout of the TM9SF4 gene aggravated the disease symptoms. In colonic epithelial cells, short hairpin RNA-mediated knockdown of TM9SF4 expression promoted inflammation and increased ER stress. In macrophages, TM9SF4 knockdown promoted M1 macrophage polarization but suppressed M2 macrophage polarization. Genetic knockout/knockdown of TM9SF4 also disrupted epithelial barrier function. Mechanistically, TM9SF4 deficiency may act through Ca2+ store depletion and cytosolic acidification to induce an ER stress increase. Furthermore, the expression level of TM9SF4 was found to be much lower in the inflamed colon regions of human ulcerative colitis patients than in normal colon samples. CONCLUSIONS: Our study identified a novel IBD-associated protein, TM9SF4, the reduced expression of which can aggravate intestinal inflammation. Deficiency of TM9SF4 increases ER stress, promotes inflammation, and impairs the intestinal epithelial barrier to aggravate IBD.

TM9SF4 is a novel regulator in lineage commitment of bone marrow mesenchymal stem cells to either osteoblasts or adipocytes.

BACKGROUND: Osteoporosis is a common bone disease in elderly population caused by imbalanced bone formation and bone resorption. Mesenchymal stem cells (MSCs) are responsible for maintaining this bone homeostasis. The phenotype of transmembrane 9 superfamily 4 (TM9SF4) knockout mice suggests a relationship between TM9SF4 proteins and bone homeostasis. But the effect of TM9SF4 in osteology has never been reported. In the present study, we investigated the function of TM9SF4 in MSC differentiation commitment, as well as its role in osteoporosis. METHODS: Primary bone marrow MSCs, isolated from TM9SF4 wildtype (TM9SF4+/+) and knockout (TM9SF4-/-) mice, were induced to differentiate into osteoblasts or adipocytes, respectively. The osteogenesis was examined by qRT-PCR detection of osteogenic markers, ALP staining and Alizarin Red S staining. The adipogenesis was tested by qRT-PCR quantification of adipogenic markers and Oil Red O staining. The cytoskeletal organization of MSCs was observed under confocal microscope. The osteoporotic model was induced by ovariectomy in TM9SF4+/+ and TM9SF4-/- mice, followed by Toluidine blue and H&E staining to assess lipid accumulation in trabecular bones, as well as micro-computed tomography scanning and immunohistochemistry staining for bone mass density assessment. The experiments on signaling pathways were conducted using qRT-PCR, Western blot and Alizarin Red S staining. RESULTS: We determined the role of TM9SF4 in MSC differentiation and found that TM9SF4-/- MSCs had higher potential to differentiate into osteoblasts and lower capability into adipocytes, without affecting osteoclastogenesis in vitro. In ovariectomy-induced osteoporotic model, TM9SF4-/- mice retained higher bone mass and less lipid accumulation in trabecular bones, indicating an important role of TM9SF4 in the regulation of osteoporosis. Mechanistically, TM9SF4-depleted cells showed elongated actin fibers, which may act through mTORC2/Akt/ß-catenin pathway to promote their commitment into osteoblasts. Furthermore, TM9SF4-depleted cells showed higher activity of canonical Wnt pathway, suggesting the participation of Wnt/ß-catenin during TM9SF4-regulated osteogenesis. CONCLUSIONS: Our study demonstrates TM9SF4 as a novel regulator for MSC lineage commitment. Depletion of TM9SF4 preferentially drives MSCs into osteoblasts instead of adipocytes. Furthermore, TM9SF4-/- mice show delayed bone loss and reduced lipid accumulation during ovariectomy-induced osteoporosis. Our results indicate TM9SF4 as a promising target for the future clinical osteoporotic treatment.

TM9SF4 levels determine sorting of transmembrane domains in the early secretory pathway.

Previous studies have shown that TM9SF4 interacts with glycine-rich transmembrane domains (TMDs) and promotes their surface localization, presumably by escorting them along the secretory pathway. Here, we delineated the role of TM9 proteins in the sorting of TMDs. Our results indicate that TM9SF4 interacts with and sorts a variety of TMDs. In human embryonic kidney (HEK) cells, a TMD carrying a positively charged residue (T-R1) or a negatively charged residue (T-D1) was localized to the endoplasmic reticulum (ER), but partially relocated to the Golgi complex upon overexpression of TM9SF4. These results show that TM9SF4 controls the sorting of TMDs at the ER-Golgi interface. Remarkably, sorting of T-R1 in HCT116 cells was different from that in HEK cells: in HCT116 cells, a substantial fraction of T-R1 was localized to the Golgi complex, and it was relocated to the ER by genetic ablation of TM9SF4. This observation indicates that TM9SF4 sorting activity differs in HEK and HCT116 cells, resulting in different sorting of TMDs in these two cell types. Although TM9SF1 associated with several TMDs, it did not visibly alter their intracellular transport in the secretory pathway and may function in other intracellular transport pathways.