Molecular Pathogenesis of Human Immunodeficiency Virus-Associated Disease of Oropharyngeal Mucosal Epithelium.

The oropharyngeal mucosal epithelia have a polarized organization, which is critical for maintaining a highly efficient barrier as well as innate immune functions. In human immunodeficiency virus (HIV)/acquired immune deficiency syndrome (AIDS) disease, the barrier and innate immune functions of the oral mucosa are impaired via a number of mechanisms. The goal of this review was to discuss the molecular mechanisms of HIV/AIDS-associated changes in the oropharyngeal mucosa and their role in promoting HIV transmission and disease pathogenesis, notably the development of opportunistic infections, including human cytomegalovirus, herpes simplex virus, and Epstein-Barr virus. In addition, the significance of adult and newborn/infant oral mucosa in HIV resistance and transmission was analyzed. HIV/AIDS-associated changes in the oropharyngeal mucosal epithelium and their role in promoting human papillomavirus-positive and negative neoplastic malignancy are also discussed

Activity-dependent regulation of miRNAs in different subcellular compartments of neurons and its implications for neuronal morphogenesis and plasticity

The activity-dependent spatiotemporal regulation of gene expression in neurons is essential for the formation and function of neuronal circuits within the brain. Recently microRNAs, a new class of post-transcriptional regulators of gene expression were implicated in the regulation of neuronal differentiation and development. Furthermore, in mature fully developed neurons, miRNAs (e.g. miR-134) were shown to be involved in the control of local protein synthesis in the vicinity of dendritic spines (Schratt et al., 2006). Activity-dependent local protein synthesis is required for synaptic plasticity, which is believed to be one of the molecular substrates of learning and memory. Nonetheless, the molecular mechanisms underlying the function and regulation of miRNAs during synaptic plasticity are poorly understood. In a previous publication from our lab, it was shown that the activity of the brain-enriched miRNA - miR-134 is regulated by brain-derived neurotrophic factor, which is released upon synaptic stimulation in neurons (Schratt et al, 2006). Interestingly, in the mouse genome this miRNA is encoded in a large miRNA cluster (miR379-410 cluster) consisting of 39 miRNAs. The expression of the miR379-410 cluster is induced upon neuronal activity by virtue of myocyte-enhancing factor 2, a transcription factor that binds to a regulatory region upstream of this cluster (Fiore et al., 2009). The transcriptional upregulation of a subset of miRNAs from the miR379-410 cluster (miR-134, -381 and -329) is necessary for activity-dependent dendritic development of rat hippocampal neurons. Furthermore, we found that the post-transcriptional regulation of the RNA-binding protein Pumilio 2 by miR-134 is essential for activity-dependent dendritogenesis. Taken together, we defined a novel MEF2-miRNA-PUM2 pathway involved in the activity-dependent regulation of dendritogenesis in primary neurons. MiR-134 localizes within dendrites of hippocampal neurons, where it can regulate the local translation of proteins important for spine structure and plasticity. However, at the beginning of this project, it was unknown how this miRNA is targeted to dendrites. I was involved in a project that aimed at identifying and characterizing the transport mechanism of miR-134 to dendrites. We found that the dendritic localization of miR-134 is mediated by the DEAH-box helicase DHX36 protein, which binds to a cis-acting element located within the loop region of the miR-134 precursor (pre-miR-134; Bicker et al., 2013). Furthermore, we showed that depletion of DHX36 increased protein levels of LIM kinase 1, a dendritically localized target of miR-134 (Schratt et al, 2006). Moreover, the depletion of DHX36 led to an increase in dendritic spine size, a similar phenotype as observed upon inhibition of miR-134 activity. In summary, we described a novel mechanism for dendritic targeting of pre-miR-134 relevant for the function of miR-134 in spine morphogenesis. Activity-dependent regulation of gene expression in the nucleus is important for the development and function of the nervous system, including synaptic plasticity and memory formation. Interestingly, several recent reports suggested that miRNAs (and/or siRNAs) might be involved in the regulation of epigenetic modifications and alternative mRNA splicing events in the nucleus of non-neuronal cells. However, whether miRNAs employ this mechanism to regulate gene expression in the neuronal nucleus was not known. A prerequisite for the study of miRNA function in the nucleus of post-mitotic neurons is the a priori knowledge of the nuclear miRNA repertoire. Therefore, using microarray and deep sequencing technologies, I identified miRNAs which are enriched in the nuclei of rat primary cortical neurons (Khudayberdiev et al. 2013; Frontiers in Mol. Neurosci, accepted for publication). Subsequently, I validated differential expression of specific nuclear-enriched miRNAs by Northern blot, quantitative real-time PCR and fluorescence in situ hybridization. By cross-comparison to published reports, I found that nuclear accumulation of miRNAs might be linked to a down-regulation of their expression during in vitro development of cortical neurons. Importantly, I found a significant overrepresentation of guanine nucleotides at the 3’ terminus of nuclear-enriched miRNA isoforms (isomiRs), suggesting the presence of neuron-specific mechanisms involved in miRNA nuclear localization. In conclusion, these results provide a starting point for future studies addressing the nuclear function of specific miRNAs and the detailed mechanisms underlying subcellular localization of miRNAs in neurons. Taken together, the results presented in my cumulative PhD thesis demonstrate that activity-dependent regulation of specific miRNAs in different subcellular neuronal compartments (dendrites, nucleus, and soma) plays an important role in neuronal morphogenesis (dendrite and spine development) and plasticity

HIV-1 Tat-induced disruption of epithelial junctions and epithelial-mesenchymal transition of oral and genital epithelial cells lead to increased invasiveness of neoplastic cells and the spread of herpes simplex virus and cytomegalovirus

Human immunodeficiency virus (HIV-1) transactivator Tat is a unique multi-functional viral protein secreted by infected cells. Although its primary function is to promote HIV-1 transcription, secreted Tat interacts with neighboring cells and induces numerous disease-associated pathological changes. Despite the substantial reduction of viral load and disease burden, Tat expression and secretion persist in people living with HIV who are undergoing treatment with highly effective combination antiretroviral therapy (cART). Tat interacts with both oral and genital epithelial cells and impairs their mucosal barrier functions, which facilitates the entry of other pathogenic viruses. Tat-mediated interactions with both human papillomavirus (HPV) -infected and HPV-negative neoplastic epithelial cells lead to epithelial-mesenchymal transition and increased invasiveness of malignant cells. Likewise, Tat-induced disruption of oral epithelial cell junctions leads to herpes simplex virus-1 (HSV-1) infection and spread via exposure of its receptor, nectin-1. HIV-1 Tat facilitates infection and spread of human cytomegalovirus (HCMV) by activating mitogen-activated protein kinases (MAPK) and promoting NF-κB signaling, both critical for the replication and production of progeny virions. HIV extracellular Tat also plays a critical role in human herpesvirus 8 (HHV8) -caused Kaposi sarcoma (KS) pathogenesis by synergizing with HHV-8 lytic proteins and promoting the proliferation, angiogenesis, and migration of endothelial cells. Collectively, these findings emphasize the critical impact of HIV-1 Tat on HIV/AIDS pathogenesis during the cART era and highlight the need for further research on the molecular mechanisms underlying Tat-mediated interactions with oral and genital mucosal epithelial cells

Economic Efficiency of Use of Water Resources in Agriculture

In this article, we discuss the economic efficiency of the use of water resources in agriculture, the use of irrigation water in agriculture, determining water consumption in irrigated agriculture and finding an effective solution to it, the efficiency of irrigation systems, and chemicalization of agriculture. we will learn about broad concepts

Release of HIV-1 sequestered in the vesicles of oral and genital mucosal epithelial cells by epithelial-lymphocyte interaction.

Oropharyngeal mucosal epithelia of fetuses/neonates/infants and the genital epithelia of adults play a critical role in HIV-1 mother-to-child transmission and sexual transmission of virus, respectively. To study the mechanisms of HIV-1 transmission through mucosal epithelium, we established polarized tonsil, cervical and foreskin epithelial cells. Analysis of HIV-1 transmission through epithelial cells showed that approximately 0.05% of initially inoculated virions transmigrated via epithelium. More than 90% of internalized virions were sequestered in the endosomes of epithelial cells, including multivesicular bodies (MVBs) and vacuoles. Intraepithelial HIV-1 remained infectious for 9 days without viral release. Release of sequestered intraepithelial HIV-1 was induced by the calcium ionophore ionomycin and by cytochalasin D, which increase intracellular calcium and disrupt the cortical actin of epithelial cells, respectively. Cocultivation of epithelial cells containing HIV-1 with activated peripheral blood mononuclear cells and CD4+ T lymphocytes led to the disruption of epithelial cortical actin and spread of virus from epithelial cells to lymphocytes. Treatment of epithelial cells with proinflammatory cytokines tumor necrosis factor-alpha and interferon gamma also induced reorganization of cortical actin and release of virus. Inhibition of MVB formation by small interfering RNA (siRNA)-mediated silencing of its critical protein hepatocyte growth factor-regulated tyrosine kinase substrate (Hrs) expression reduced viral sequestration in epithelial cells and its transmission from epithelial cells to lymphocytes by ~60-70%. Furthermore, inhibition of vacuole formation of epithelial cells by siRNA-inactivated rabankyrin-5 expression also significantly reduced HIV-1 sequestration in epithelial cells and spread of virus from epithelial cells to lymphocytes. Interaction of the intercellular adhesion molecule-1 of epithelial cells with the function-associated antigen-1 of lymphocytes was important for inducing the release of sequestered HIV-1 from epithelial cells and facilitating cell-to-cell spread of virus from epithelial cells to lymphocytes. This mechanism may serve as a pathway of HIV-1 mucosal transmission