High expression of TMEM33 predicts poor prognosis and promotes cell proliferation in cervical cancer

Background As one of the most common cancer among women worldwide, the prognosis of patients with advanced cervical cancer remains unsatisfactory. A study indicated that transmembrane protein 33 (TMEM33) was implicated in tumor recurrence, while its role in cervical cancer has not been elucidated. Methods TMEM33 expression in cervical squamous cell carcinoma and endocervical adenocarcinoma (CESC) was primarily screened in The Cancer Genome Altas (TCGA), and further validated in Gene Expression Omnibus (GEO) database. The Kaplan–Meier plotter analysis and Cox regression were constructed to evaluate the prognostic value of TMEM33 in CESC. Functional enrichment analysis was performed with GO, KEGG and GSEA tools. Protein-protein interaction analysis and correlated gene networks were conducted using STING and GEPIA2 websites, respectively. The expression of TMEM33 in cervical cancer cells were examined by immunoblotting and RT-qPCR. Finally, CCK-8 assay and colony formation assay were performed to investigate the role of TMEM33 in cervical cancer cell proliferation. Results TMEM33 expression was significantly elevated in CESC compared with normal tissues. High expression of TMEM33 was associated with poor prognostic clinical characteristics in CESC patients. KM-plotter analysis revealed that patients with increased TMEM33 had shorter overall survival (OS), progress free interval (PFI), and disease specific survival (DSS). Moreover, Multivariate Cox analysis further confirmed that high TMEM33 expression was an independent risk factor for OS in patients with CESC. TMEM33 was associated with immune cell infiltration, and its expression was correlated with tumorigenesis-related genes RNF4, OCIAD1, TMED5, DHX15, MED28 and LETM1. More importantly, knockdown of TMEM33 in cervical cancer cells decreased the expression of those genes and inhibited cell proliferation. Conclusions Increased TMEM33 in cervical cancer can serve as an independent prognostic marker and might play a role in tumorigenesis by promoting cell proliferation.

A mutation in transmembrane protein 135 impairs lipid metabolism in mouse eyecups

Aging is a significant factor in the development of age-related diseases but how aging disrupts cellular homeostasis to cause age-related retinal disease is unknown. Here, we further our studies on transmembrane protein 135 (Tmem135), a gene involved in retinal aging, by examining the transcriptomic profiles of wild-type, heterozygous and homozygous Tmem135 mutant posterior eyecup samples through RNA sequencing (RNA-Seq). We found significant gene expression changes in both heterozygous and homozygous Tmem135 mutant mouse eyecups that correlate with visual function deficits. Further analysis revealed that expression of many genes involved in lipid metabolism are changed due to the Tmem135 mutation. Consistent with these changes, we found increased lipid accumulation in mutant Tmem135 eyecup samples. Since mutant Tmem135 mice have similar ocular pathologies as human age-related macular degeneration (AMD) eyes, we compared our homozygous Tmem135 mutant eyecup RNA-Seq dataset with transcriptomic datasets of human AMD donor eyes. We found similar changes in genes involved in lipid metabolism between the homozygous Tmem135 mutant eyecups and AMD donor eyes. Our study suggests that the Tmem135 mutation affects lipid metabolism as similarly observed in human AMD eyes, thus Tmem135 mutant mice can serve as a good model for the role of dysregulated lipid metabolism in AMD.

Secreted Trimeric Chikungunya Virus Spikes from Insect Cells: Production, Purification, and Glycosylation Status

Chikungunya virus (CHIKV) is a rapidly emerging mosquito-borne virus that causes a severe febrile illness with long-lasting arthralgia in humans. As there is no vaccine to protect humans and limit CHIKV epidemics, the virus continues to be a global public health concern. The CHIKV envelope glycoproteins E1 and E2 are important immunogens; therefore, the aim of this study is to produce trimeric CHIKV spikes in insect cells using the baculovirus expression system. The CHIKV E1 and E2 ectodomains were covalently coupled by a flexible linker that replaces the 6K transmembrane protein. The C-terminal E1 transmembrane was replaced by a Strep-tag II for the purification of secreted spikes from the culture fluid. After production in Sf9 suspension cells (product yields of 5.8–7.6 mg/L), the CHIKV spikes were purified by Strep-Tactin affinity chromatography, which successfully cleared the co-produced baculoviruses. Bis(sulfosuccinimidyl)suberate cross-linking demonstrated that the spikes are secreted as trimers. PNGase F treatment showed that the spikes are glycosylated. LC–MS/MS-based glycoproteomic analysis confirmed the glycosylation and revealed that the majority are of the mannose- or hybrid-type N-glycans and <2% have complex-type N-glycans. The LC –MS/MS analysis also revealed three O-glycosylation sites in E1. In conclusion, the trimeric, glycosylated CHIKV spikes have been successfully produced in insect cells and are now available for vaccination studies.

Prom1 and Notch regulate ciliary length and dynamics in multiciliated cells of the airway epithelium

Differences in ciliary morphology and dynamics among multiciliated cells of the respiratory tract have been well reported and known to contribute to efficient mucociliary clearance. Nevertheless, little is known about how phenotypic differences among multiciliated cells are established in the mammalian lung. Here we show that Prominin-1 (Prom1), a transmembrane protein widely used as stem cell and tumor-initiating marker, is crucial to this process. During airway differentiation, Prom1 becomes restricted to multiciliated cells, where it is expressed at distinct levels along the proximal-distal axis of the airways and in the adult airway epithelium in vitro. We found that Prom1 is induced by Notch in post-specified multiciliated cells and that Notch inactivation abolishes the gradients of Prom1 in the developing airways and in differentiating organotypic cultures. Prom1 was not required for multicilia formation and when inactivated resulted in longer cilia, which remained functional but beating at a lower frequency. Disruption of Notch resulted in opposite effects and suggested that Notch fine-tunes Prom1 levels to regulate the multiciliated cell phenotype and generate diversity among these cells in the respiratory tract. By controlling these features, this mechanism contributes to the innate defense of the lung against environmental agents and prevent pulmonary disease.

LRRTM3 regulates activity-dependent synchronization of synapse properties in topographically connected hippocampal neural circuits

Synaptic cell-adhesion molecules (CAMs) organize the architecture and properties of neural circuits. However, whether synaptic CAMs are involved in activity-dependent remodeling of specific neural circuits is incompletely understood. Leucine-rich repeat transmembrane protein 3 (LRRTM3) is required for the excitatory synapse development of hippocampal dentate gyrus (DG) granule neurons. Here, we report that Lrrtm3-deficient mice exhibit selective reductions in excitatory synapse density and synaptic strength in projections involving the medial entorhinal cortex (MEC) and DG granule neurons, accompanied by increased neurotransmitter release and decreased excitability of granule neurons. LRRTM3 deletion significantly reduced excitatory synaptic innervation of hippocampal mossy fibers (Mf) of DG granule neurons onto thorny excrescences in hippocampal CA3 neurons. Moreover, LRRTM3 loss in DG neurons significantly decreased mossy fiber long-term potentiation (Mf-LTP). Remarkably, silencing MEC–DG circuits protected against the decrease in the excitatory synaptic inputs onto DG and CA3 neurons, excitability of DG granule neurons, and Mf-LTP in Lrrtm3-deficient mice. These results suggest that LRRTM3 may be a critical factor in activity-dependent synchronization of the topography of MEC–DG–CA3 excitatory synaptic connections. Collectively, our data propose that LRRTM3 shapes the target-specific structural and functional properties of specific hippocampal circuits.

Calcium Signaling Pathway Is Involved in the Shedding of ACE2 Catalytic Ectodomain: New Insights for Clinical and Therapeutic Applications of ACE2 for COVID-19

The angiotensin-converting enzyme 2 (ACE2) is a type I integral membrane that exists in two forms: the first is a transmembrane protein; the second is a soluble catalytic ectodomain of ACE2. The catalytic ectodomain of ACE2 undergoes shedding by a disintegrin and metalloproteinase domain-containing protein 17 (ADAM17), in which calmodulin mediates the calcium signaling pathway that is involved in ACE2 release, resulting in a soluble catalytic ectodomain of ACE2 that can be measured as soluble ACE2 plasma activity. The shedding of the ACE2 catalytic ectodomain plays a role in cardiac remodeling and endothelial dysfunction and is a predictor of all-cause mortality, including cardiovascular mortality. Moreover, considerable evidence supports that the ACE2 catalytic ectodomain is an essential entry receptor for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. Additionally, endotoxins and the pro-inflammatory cytokines interleukin (IL)-1β and tumor necrosis factor-alpha (TNFα) all enhanced soluble catalytic ectodomain ACE2 shedding from the airway epithelia, suggesting that the shedding of ACE2 may represent a mechanism by which viral entry and infection may be controlled such as some types of betacoronavirus. In this regard, ACE2 plays an important role in inflammation and thrombotic response, and its down-regulation may aggravate COVID-19 via the renin-angiotensin system, including by promoting pathological changes in lung injury. Soluble forms of ACE2 have recently been shown to inhibit SARS-CoV-2 infection. Furthermore, given that vitamin D enhanced the shedding of ACE2, some studies reported that vitamin D treatment is associated with prognosis improvement in COVID-19. This is an updated review on the evidence, clinical, and therapeutic applications of ACE2 for COVID-19.

Role and Involvement of TENM4 and miR-708 in Breast Cancer Development and Therapy

Teneurin 4 (TENM4) is a transmembrane protein that is codified by the ODZ4 gene and is involved in nervous system development, neurite outgrowth, and neuronal differentiation. In line with its involvement in the nervous system, TENM4 has also been implicated in several mental disorders such as bipolar disorder, schizophrenia, and autism. TENM4 mutations and rearrangements have recently been identified in a number of tumors. This, combined with impaired expression in tumors, suggests that it may potentially be involved in tumorigenesis. Most of the TENM4 mutations that are observed in tumors occur in breast cancer, in which TENM4 plays a role in cells’ migration and stemness. However, the functional role that TENM4 plays in breast cancer still needs to be better evaluated, and further studies are required to better understand the involvement of TENM4 in breast cancer progression. Herein, we review the currently available data for TENM4′s role in breast cancer and propose its use as both a novel target with which to ameliorate patient prognosis and as a potential biomarker. Moreover, we also report data on the tumorigenic role of miR-708 deregulation and the possible use of this miRNA as a novel therapeutic molecule, as miR-708 is spliced out from TENM4 mRNA.

Neuropilin-1, a myeloid cell-specific protein, is an inhibitor of HIV-1 infectivity

Myeloid lineage cells such as macrophages and dendritic cells (DCs), targeted by HIV-1, are important vehicles for virus dissemination through the body as well as viral reservoirs. Compared to activated lymphocytes, myeloid cells are collectively more resistant to HIV-1 infection. Here we report that NRP-1, encoding transmembrane protein neuropilin-1, is highly expressed in macrophages and DCs but not CD4+ T cells, serving as an anti-HIV factor to inhibit the infectivity of HIV-1 progeny virions. Silencing NRP-1 enhanced the transmission of HIV-1 in macrophages and DCs significantly and increased the infectivity of the virions produced by these cells. We further demonstrated that NRP-1 was packaged into the progeny virions to inhibit their ability to attach to target cells, thus reducing the infectivity of the virions. These data indicate that NRP-1 is a newly identified antiviral protein highly produced in both macrophages and DCs that inhibit HIV-1 infectivity; thus, NRP-1 may be a novel therapeutic strategy for the treatment of HIV-1 infection.