Biology subjects

Zurita, J.

Publications and source records attributed to Zurita, J..

5 recordsLinked to original sources

bioRxiv2022

Glutarate regulates T cell function and metabolism

T cell function is influenced by several metabolites; some acting through enzymatic inhibition of -KG-dependent dioxygenases (KGDDs), others, through post-translational modification of lysines in important targets. We show here that glutarate, a product of amino acid catabolism, has the capacity to do both, with effects on T cell function and differentiation. Glutarate exerts those effects through KGDD inhibition and through direct regulation of T cell metabolism via post-translational modification of the pyruvate dehydrogenase E2 subunit. Diethyl-glutarate, a cell-permeable form of glutarate, alters CD8+ T cell differentiation and increases cytotoxicity against target cells. In vivo administration of the compound reduces tumor growth and is correlated with increased levels of both peripheral and intratumoral cytotoxic CD8+ T cells. These results demonstrate that glutarate regulates both T cell metabolism and differentiation, with a potential role in the improvement of T cell immunotherapy.

immunology↗

bioRxiv2021

Ku70 suppresses alternative end-joining in G1-arrested progenitor B cells

Classical nonhomologous end-joining (C-NHEJ) repairs DNA double-stranded breaks (DSBs) throughout interphase but predominates in G1-phase when homologous recombination is unavailable. Complexes containing the Ku70/80 ("Ku") and XRCC4/Ligase IV (Lig4) core C-NHEJ factors are required, respectively, for sensing and joining DSBs. While XRCC4/Ligase IV are absolutely required for joining RAG1/2-endonucease ("RAG")-initiated DSBs during V(D)J recombination in G1-phase progenitor lymphocytes, cycling cells deficient for XRCC4/Ligase IV also can join chromosomal DSBs by alternative end-joining (A-EJ) pathways. Restriction of V(D)J recombination by XRCC4/Ligase IV-mediated joining has been attributed to RAG shepherding V(D)J DSBs exclusively into the C-NHEJ pathway. Here, we report that A-EJ of DSB ends generated by RAG1/2, Cas9:gRNA and Zinc finger endonucleases in Lig4-deficient G1-arrested progenitor B cell lines is suppressed by Ku. Thus, while diverse DSBs remain largely as free broken ends in Lig4-deficient G1-arrested progenitor B cells, deletion of Ku70 increases DSB rejoining and translocation levels to those observed in Ku70-deficient counterparts. Correspondingly, while RAG-initiated V(D)J DSB joining is abrogated in Lig4-deficient G1-arrested progenitor B cell lines, joining of RAG-generated DSBs in Ku70-deficient and Ku70/Lig4 double-deficient lines occurs through a translocation-like A-EJ mechanism. Thus, in G1-arrested, Lig4-deficient progenitor B cells are functionally end-joining suppressed due to Ku-dependent blockage of A-EJ, potentially, in association with G1-phase down-regulation of Ligase1. Finally, we suggest that differential impacts of Ku-deficiency versus Lig4-deficiency on V(D)J recombination, neuronal apoptosis, and embryonic development results from Ku-mediated inhibition of A-EJ in the G1 cell cycle phase in Lig4-defcient developing lymphocyte and neuronal cells. Significance StatementAlternative end-joining (A-EJ) is implicated in oncogenic translocations and mediating DNA double-strand break (DSB) repair in cycling cells when classical nonhomologous endjoining (C-NHEJ) factors of the C-NHEJ Ligase complex are absent. However, V(D)J recombination-associated DSBs that occur in G1 cell cycle-phase progenitor lymphocytes are joined exclusively by the C-NHEJ pathway. Until now, however, the overall mechanisms that join general DSBs in G1-phase progenitor B cells had not been fully elucidated. Here, we report that Ku, a core C-NHEJ double-strand break recognition complex, directs repair of a variety of different targeted DSBs towards C-NHEJ and suppresses A-EJ in G1-phase cells. We suggest this Ku activity explains how Ku-deficiency can rescue the neuronal development and embryonic lethality phenotype of Ligase 4-deficient mice.

genetics↗

bioRxiv2019

Induction of Recurrent Break Cluster Genes in Neural Progenitor Cells Differentiated from Embryonic Stem Cells In Culture

Mild replication stress enhances appearance of dozens of robust recurrent genomic break clusters, termed RDCs, in cultured primary mouse neural stem and progenitor cells (NSPCs). Robust RDCs occur within genes ("RDC-genes") that are long and have roles in neural cell communications and/or have been implicated in neuropsychiatric diseases or cancer. We sought to develop an in vitro approach to determine whether specific RDC formation is associated with neural development. For this purpose, we adapted a system to induce neural progenitor cell (NPC) development from mouse embryonic stem cell (ESC) lines deficient for XRCC4 plus p53, a genotype that enhances DNA double-strand break (DSB) persistence to enhance detection. We tested for RDCs by our genome wide DSB identification approach that captures DSBs genome-wide via their ability to join to specific genomic Cas9/sgRNA-generated bait DSBs. In XRCC4/p53-deficient ES cells, we detected 7 RDCs, which were in genes, with two RDCs being robust. In contrast, in NPCs derived from these ES cell lines, we detected 29 RDCs, a large fraction of which were robust and associated with long, transcribed neural genes that were also robust RDC-genes in primary NSPCs. These studies suggest that many RDCs present in NSPCs are developmentally influenced to occur in this cell type and indicate that induced development of NPCs from ES cells provides an approach to rapidly elucidate mechanistic aspects of NPC RDC formation. SIGNIFICANCE STATEMENTWe previously discovered a set of long neural genes susceptible to frequent DNA breaks in primary mouse brain progenitor cells. We termed these genes RDC-genes. RDC-gene breakage during brain development might alter neural gene function and contribute to neurological diseases and brain cancer. To provide an approach to characterize the unknown mechanism of neural RDC-gene breakage, we asked whether RDC-genes appear in neural progenitors differentiated from embryonic stem cells in culture. Indeed, robust RDC-genes appeared in neural progenitors differentiated in culture and many overlapped with robust RDC-genes in primary brain progenitors. These studies indicate that in vitro development of neural progenitors provides a model system for elucidating how RDC-genes are formed.

neuroscience↗

bioRxiv2018

Characterization of cefotaxime resistant Escherichia coli isolated from broiler farms in Ecuador

Antimicrobial resistance (AR) is a worldwide concern. Up to a 160% increase in antibiotic usage in food animals is expected in Latin American countries. The poultry industry is an increasingly important segment of food production and contributor to AR. The objective of this study was to evaluate the prevalence, AR patterns and the characterization of relevant resistance genes in Extended Spectrum {beta}-lactamases (ESBL) and AmpC E. coli from large poultry farms in Ecuador. Sampling was performed from June 2013 to July 2014 in 6 slaughterhouses that slaughter broilers from 115 farms totaling 384 flocks. Each sample of collected caeca was streaked onto TBX agar supplemented with cefotaxime (3 mg/l). In total, 176 isolates were analyzed for antimicrobial resistance patterns by the disk diffusion method and for blaCTX-M, blaTEM, blaCMY, blaSHV, blaKPC, and mcr-1 by PCR and sequencing. ESBL and AmpC E. coli were found in 362 flocks (94.3%) from 112 farms (97.4%). We found that 98.3% of the isolates were multi-resistant to antibiotics. Low resistance was observed for ertapenem and nitrofurantoin. The most prevalent ESBL genes were the blaCTX-M (90.9%) blaCTX-M-65, blaCTX-M-55 and blaCTX-M-3 alleles. Most of the AmpC strains presented the blaCMY-2 gene. Three isolates showed the mcr-1 gene. Poultry production systems represent a hotspot for antimicrobial resistance in Ecuador, possibly mediated by the extensive use of antibiotics. Monitoring this sector in national and regional plans of antimicrobial resistance surveillance should therefore be considered.

microbiology↗

bioRxiv2018

Diverse Escherichia coli lineages, from domestic animals and humans in a household, carry colistin resistance gene mcr-1 in Ecuador

The aim of this study was to investigate the presence of Escherichia coli carrying mcr-1 gene in domestic animals close to a child who suffered a peritoneal infection by a mcr-1 positive E. coli. Rectal or cloacal swabs and fecal samples from domestic animals were plated on selective media to isolate colistin-resistant E. coli and isolates were submitted to detection of mcr-1 gene, pulsed field gel electrophoresis (PFGE), multi-locus sequence typing (MLST), replicon typing and S1-PFGE. Four mcr-1 positive E. coli isolates (from chicken, turkey and dog) were recovered. No shared PFGE pattern or MLST sequence type were observed among isolates. A 60Kb IncI1{gamma} mcr-1-carrying plasmid was detected in all isolates. Our results suggest that mcr-1 gene was horizontally disseminated amongst different lineages of E. coli from domestic animals in the childs household.\n\nImportanceHorizontally transferable colistin resistance (mcr-1 gene) is thought to have originated in domestic animals and transferred to humans through meat and dairy products. In the present report we show evidence that the mcr-1 gene could be transferred to different E. coli strains colonizing different hosts (humans and pets) in the same household.

microbiology↗