Emergence and spread of a B.1.1.28-derived lineage with Q675H and Q677H Spike mutations in Uruguay

Uruguay was able to control the viral dissemination during the first nine months of the SARS-CoV-2 pandemic. Unfortunately, towards the end of 2020, the number of daily new cases exponentially increased. Herein we analyzed the country-wide genetic diversity of SARS-CoV-2 between November, 2020 and April, 2021. Our findings identified that the most prevalent viral variant during late 2020 was a B.1.1.28 sublineage carrying mutations Q675H+Q677H in the viral Spike, now designated as lineage P.6. This new lineage P.6 probably arose around November 2020, in Montevideo, Uruguays capital department and rapidly spread to other Uruguayan departments, with evidence of further local transmission clusters, also spread sporadically to the USA and Spain. The Q675H and Q677H mutations are in the proximity of the polybasic cleavage site at the S1/S2 boundary and also arose independently in many SARS-CoV-2 lineages circulating worldwide. Although the lineage P.6 was replaced by the Variant of Concern (VOC) P.1 as the predominant viral strain in Uruguay since April 2021, the monitoring of the concurrent emergence of Q675H+Q677H in VOCs should be of worldwide interest.

Roles of the ubiquitin ligase CUL4B and ADP-ribosyltransferase TiPARP in TCDD-induced nuclear export and proteasomal degradation of the transcription factor AHR.

The aryl hydrocarbon receptor (AHR) is a transcription factor activated by exogenous halogenated polycyclic aromatic hydrocarbon compounds, including the environmental toxin TCDD, 2,3,7,8-tetrachlorodibenzo-p-dioxin, and naturally occurring dietary and endogenous compounds. The activated AHR enhances transcription of specific genes including phase I and phase II metabolism enzymes and other targets genes such as the TCDD-inducible poly(ADP-ribose) polymerase (TiPARP). The regulation of AHR activation is a dynamic process: immediately after transcriptional activation of the AHR by TCDD, the AHR is exported from the nucleus to the cytoplasm where it is subjected to proteasomal degradation. However, the mechanisms regulating AHR degradation are not well understood. Here, we studied the role of two enzymes reported to enhance AHR breakdown: the cullin 4B (CUL4B)AHR complex, an E3 ubiquitin ligase that targets the AHR and other proteins for ubiquitination, and TiPARP, which targets proteins for ADP-ribosylation, a posttranslational modification that can increase susceptibility to degradation. Using a WT mouse embryonic fibroblast (MEF) cell line and an MEF cell line in which CUL4B has been deleted (MEFCul4b-null), we discovered that loss of CUL4B partially prevented AHR degradation after TCDD exposure, while knocking down TiPARP in MEFCul4b-null cells completely abolished AHR degradation upon TCDD treatment. Increased TCDD-activated AHR protein levels in MEFCul4b-null and MEFCul4b-null cells in which TiPARP was knocked down led to enhanced AHR transcriptional activity, indicating that CUL4B and TiPARP restrain AHR action. This study reveals a novel function of TiPARP in controlling TCDD-activated AHR nuclear export and subsequent proteasomal degradation.

Implementation of a qPCR assay coupled with genomic surveillance for real-time monitoring of SARS-CoV-2 variants of concern.

We developed a genomic surveillance program for real-time monitoring of SARS-CoV-2 variants of concern in Uruguay. Here, we present the first results, including the proposed qPCR-VOC method, the general workflow and the report of the introduction and community transmission of the VOC P.1 in Uruguay in multiple independent events.