Loss of the miR-21 allele elevates the expression of its target genes and reduces tumorigenesis.

MicroRNA 21 (miR-21) is overexpressed in virtually all types of carcinomas and various types of hematological malignancies. To determine whether miR-21 promotes tumor development in vivo, we knocked out the miR-21 allele in mice. In response to the 7,12-dimethylbenz[a]anthracene (DMBA)/12-O-tetradecanoylphorbol-13-acetate mouse skin carcinogenesis protocol, miR-21-null mice showed a significant reduction in papilloma formation compared with wild-type mice. We revealed that cellular apoptosis was elevated and cell proliferation was decreased in mice deficient of miR-21 compared to wild-type animals. In addition, we found that a large number of validated or predicted miR-21 target genes were up-regulated in miR-21-null keratinocytes, which are precursor cells to skin papillomas. Specifically, up-regulation of Spry1, Pten, and Pdcd4 when miR-21 was ablated coincided with reduced phosphorylation of ERK, AKT, and JNK, three major downstream effectors of Ras activation that plays a predominant role in DMBA-initiated skin carcinogenesis. These results provide in vivo evidence that miR-21 exerts its oncogenic function through negatively regulating its target genes.

Interaction of the oncogenic miR-21 microRNA and the p53 tumor suppressor pathway.

MicroRNA-21 (miR-21) is overexpressed virtually in all human cancers and displays oncogenic activity in a transgenic murine model. Similarly, the p53 tumor suppressor gene is the most frequently mutated gene in human cancer, and its loss or mutation leads to tumor formation in mice. To ascertain the role of miR-21 in the p53 pathway in vivo and to characterize their interaction in tumorigenesis, we intercrossed the miR-21 (-/-) and Trp53 (-/-) mice. We found that Trp53 (-/-) miR-21 (-/-) mice develop tumors at a slightly later age, yet show a similar tumor spectrum and survival curve as Trp53 (-/-) mice. When subjected to genotoxic agents, tissues from Trp53 (-/-) miR-21 (-/-) mice have a higher percentage of apoptotic cells. We extracted mouse embryonic fibroblast cells (MEFs) to examine the impact of miR-21 loss on p53-regulated cellular processes in Trp53 (-/-) cells. Higher cellular apoptosis and senescence were found in Trp53 (-/-) miR-21 (-/-) MEFs than in Trp53 (-/-) MEFs. In addition, loss of miR-21 sensitizes transformed Trp53 (-/-) cells to DNA damage-induced apoptosis through elevation of Pten expression. These data suggest that inhibition of miR-21 would be beneficial in apoptosis-inducing cancer therapies directed against p53-deficient tumors.

SARS-CoV-2 infection of the oral cavity and saliva.

Despite signs of infection-including taste loss, dry mouth and mucosal lesions such as ulcerations, enanthema and macules-the involvement of the oral cavity in coronavirus disease 2019 (COVID-19) is poorly understood. To address this, we generated and analyzed two single-cell RNA sequencing datasets of the human minor salivary glands and gingiva (9 samples, 13,824 cells), identifying 50 cell clusters. Using integrated cell normalization and annotation, we classified 34 unique cell subpopulations between glands and gingiva. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) viral entry factors such as ACE2 and TMPRSS members were broadly enriched in epithelial cells of the glands and oral mucosae. Using orthogonal RNA and protein expression assessments, we confirmed SARS-CoV-2 infection in the glands and mucosae. Saliva from SARS-CoV-2-infected individuals harbored epithelial cells exhibiting ACE2 and TMPRSS expression and sustained SARS-CoV-2 infection. Acellular and cellular salivary fractions from asymptomatic individuals were found to transmit SARS-CoV-2 ex vivo. Matched nasopharyngeal and saliva samples displayed distinct viral shedding dynamics, and salivary viral burden correlated with COVID-19 symptoms, including taste loss. Upon recovery, this asymptomatic cohort exhibited sustained salivary IgG antibodies against SARS-CoV-2. Collectively, these data show that the oral cavity is an important site for SARS-CoV-2 infection and implicate saliva as a potential route of SARS-CoV-2 transmission.

Integrated Single-Cell Atlases Reveal an Oral SARS-CoV-2 Infection and Transmission Axis.

Despite signs of infection, the involvement of the oral cavity in COVID-19 is poorly understood. To address this, single-cell RNA sequencing data-sets were integrated from human minor salivary glands and gingiva to identify 11 epithelial, 7 mesenchymal, and 15 immune cell clusters. Analysis of SARS-CoV-2 viral entry factor expression showed enrichment in epithelia including the ducts and acini of the salivary glands and the suprabasal cells of the mucosae. COVID-19 autopsy tissues confirmed in vivo SARS-CoV-2 infection in the salivary glands and mucosa. Saliva from SARS-CoV-2-infected individuals harbored epithelial cells exhibiting ACE2 expression and SARS-CoV-2 RNA. Matched nasopharyngeal and saliva samples found distinct viral shedding dynamics and viral burden in saliva correlated with COVID-19 symptoms including taste loss. Upon recovery, this cohort exhibited salivary antibodies against SARS-CoV-2 proteins. Collectively, the oral cavity represents a robust site for COVID-19 infection and implicates saliva in viral transmission.

miR-21 and let-7 in the Ras and NF-κB pathways.

miR-21 and let-7 are two of the most studied microRNAs (miRNAs), as the former is the most frequently upregulated and the latter is the most frequently down-regulated in cancer. In this short essay, we examine the convergence of miR-21 and let-7 in two major cancer pathways: Ras and NF-κB. miR-21 suppresses multiple targets to enhance the oncogenic action of Ras, while let-7 is a direct negative regulator of the Ras gene family. let-7 is proposed to repress the activation of NF-κB through down-regulation of Ras and IL-6, while miR-21 is demonstrated to enhance NF-κB activation by down-regulating Pten and thereby increasing the activity of Akt, a kinase known to promote the NF-κB pathway. Therefore, miR-21 and let-7 contravene each other in cancer and simultaneous targeting of both is an attractive therapeutic strategy against cancers exhibiting both Ras mutations and constitutive NF-κB activation.