Pro-CRISPR PcrIIC1-associated Cas9 system for enhanced bacterial immunity.

The CRISPR system is an adaptive immune system found in prokaryotes that defends host cells against the invasion of foreign DNA1. As part of the ongoing struggle between phages and the bacterial immune system, the CRISPR system has evolved into various types, each with distinct functionalities2. Type II Cas9 is the most extensively studied of these systems and has diverse subtypes. It remains uncertain whether members of this family can evolve additional mechanisms to counter viral invasions3,4. Here we identify 2,062 complete Cas9 loci, predict the structures of their associated proteins and reveal three structural growth trajectories for type II-C Cas9. We found that novel associated genes (NAGs) tended to be present within the loci of larger II-C Cas9s. Further investigation revealed that CbCas9 from Chryseobacterium species contains a novel ß-REC2 domain, and forms a heterotetrameric complex with an NAG-encoded CRISPR-Cas-system-promoting (pro-CRISPR) protein of II-C Cas9 (PcrIIC1). The CbCas9-PcrIIC1 complex exhibits enhanced DNA binding and cleavage activity, broader compatibility for protospacer adjacent motif sequences, increased tolerance for mismatches and improved anti-phage immunity, compared with stand-alone CbCas9. Overall, our work sheds light on the diversity and 'growth evolutionary' trajectories of II-C Cas9 proteins at the structural level, and identifies many NAGs-such as PcrIIC1, which serves as a pro-CRISPR factor to enhance CRISPR-mediated immunity.

The CRISPR effector Cam1 mediates membrane depolarization for phage defence.

Prokaryotic type III CRISPR-Cas systems provide immunity against viruses and plasmids using CRISPR-associated Rossman fold (CARF) protein effectors1-5. Recognition of transcripts of these invaders with sequences that are complementary to CRISPR RNA guides leads to the production of cyclic oligoadenylate second messengers, which bind CARF domains and trigger the activity of an effector domain6,7. Whereas most effectors degrade host and invader nucleic acids, some are predicted to contain transmembrane helices without an enzymatic function. Whether and how these CARF-transmembrane helix fusion proteins facilitate the type III CRISPR-Cas immune response remains unknown. Here we investigate the role of cyclic oligoadenylate-activated membrane protein 1 (Cam1) during type III CRISPR immunity. Structural and biochemical analyses reveal that the CARF domains of a Cam1 dimer bind cyclic tetra-adenylate second messengers. In vivo, Cam1 localizes to the membrane, is predicted to form a tetrameric transmembrane pore, and provides defence against viral infection through the induction of membrane depolarization and growth arrest. These results reveal that CRISPR immunity does not always operate through the degradation of nucleic acids, but is instead mediated via a wider range of cellular responses.

Sexo Masculino e Hipertensão Arterial São Preditores de Placa à Angiotomografia das Coronárias / Male Gender and Arterial Hypertension are Plaque Predictors at Coronary Computed Tomography Angiography

Background: Systemic Arterial Hypertension (SAH) is one of the main risk factors for Coronary Artery Disease (CAD), in addition to male gender. Differences in coronary artery lesions between hypertensive and normotensive individuals of both genders at the Coronary Computed Tomography Angiography (CCTA) have not been clearly determined. Objective: To Investigate the calcium score (CS), CAD extent and characteristics of coronary plaques at CCTA in men and women with and without SAH. Methods: Prospective cross-sectional study of 509 patients undergoing CCTA for CAD diagnosis and risk stratification, from November 2011 to December 2012, at Instituto de Cardiologia Dante Pazzanese. Individuals were stratified according to gender and subdivided according to the presence (HT +) or absence (HT-) of SAH. Results: HT+ women were older (62.3 ± 10.2 vs 57.8 ± 12.8, p = 0.01). As for the assessment of CAD extent, the HT+ individuals of both genders had significant CAD, although multivessel disease is more frequent in HT + men. The regression analysis for significant CAD showed that age and male gender were the determinant factors of multivessel disease and CS ≥ 100. Plaque type analysis showed that SAH was a predictive risk factor for partially calcified plaques (OR = 3.9). Conclusion: Hypertensive men had multivessel disease more often than women. Male gender was a determinant factor of significant CAD, multivessel disease, CS ≥ 100 and calcified and partially calcified plaques, whereas SAH was predictive of partially calcified plaques. .

Fundamento: Doença Arterial Coronariana (DAC) tem como um dos principais fatores de risco a Hipertensão Arterial Sistêmica (HAS), além do sexo masculino. As diferenças nas lesões de artérias coronarianas entre hipertensos e normotensos de ambos os sexos pela Angiotomografia Computadorizada Coronariana (ATC) ainda não foram claramente determinadas. Objetivo: Investigar Escore de Cálcio (EC), extensão da DAC e as características de placas coronarianas à ATC em homens e mulheres com e sem HAS. Métodos: Estudo transversal prospectivo com 509 pacientes submetidos a ATC para diagnóstico e estratificação de risco de DAC, de novembro de 2011 a dezembro de 2012, no Instituto Dante Pazzanese de Cardiologia. Foram estratificados segundo o gênero e subdivididos de acordo com a presença (HT+) ou ausência (HT-) de HAS. Resultados: As mulheres HT+ apresentaram maior idade (62,3 ± 10,2 vs 57,8 ± 12,8, p = 0,01). Quanto à avaliação de extensão da DAC, os HT+ de ambos os sexos apresentaram DAC significativa, porém a doença multiarterial é mais frequente nos homens HT+. A análise da regressão para DAC significativa mostrou que idade e sexo masculino foram fatores determinantes de doença multiarterial e EC ≥ 100. Já a análise para os tipos de placas mostrou que a HAS se comportou como fator de risco preditor para placas parcialmente calcificadas (OR = 3,9). Conclusão: Homens hipertensos apresentaram mais frequentemente doença multiarterial. O sexo masculino foi fator determinante de DAC significativa, doença multiarterial, EC ≥ 100 e placas calcificadas e parcialmente calcificadas. Enquanto a HAS foi preditora de placas parcialmente calcificadas. .