Active and adaptive Legionella CRISPR‐Cas reveals a recurrent challenge to the pathogen

Clustered regularly interspaced short palindromic repeats with CRISPR‐associated gene (CRISPR‐Cas) systems are widely recognized as critical genome defense systems that protect microbes from external threats such as bacteriophage infection. Several isolates of the intracellular pathogen Legionella pneumophila possess multiple CRISPR‐Cas systems (type I‐C, type I‐F and type II‐B), yet the targets of these systems remain unknown. With the recent observation that at least one of these systems (II‐B) plays a non‐canonical role in supporting intracellular replication, the possibility remained that these systems are vestigial genome defense systems co‐opted for other purposes. Our data indicate that this is not the case. Using an established plasmid transformation assay, we demonstrate that type I‐C, I‐F and II‐B CRISPR‐Cas provide protection against spacer targets. We observe efficient laboratory acquisition of new spacers under ‘priming’ conditions, in which initially incomplete target elimination leads to the generation of new spacers and ultimate loss of the invasive DNA. Critically, we identify the first known target of L. pneumophila CRISPR‐Cas: a 30 kb episome of unknown function whose interbacterial transfer is guarded against by CRISPR‐Cas. We provide evidence that the element can subvert CRISPR‐Cas by mutating its targeted sequences – but that primed spacer acquisition may limit this mechanism of escape. Rather than generally impinging on bacterial fitness, this element drives a host specialization event – with improved fitness in Acanthamoeba but a reduced ability to replicate in other hosts and conditions. These observations add to a growing body of evidence that host range restriction can serve as an existential threat to L. pneumophila in the wild.     

Abscisic acid deficiency increases defence responses against Myzus persicae in Arabidopsis

Comparison of Arabidopsis thaliana (Arabidopsis) gene expression induced by Myzus persicae (green peach aphid) feeding, aphid saliva infiltration and abscisic acid (ABA) treatment showed a significant positive correlation. In particular, ABA‐regulated genes are over‐represented among genes that are induced by M. persicae saliva infiltration into Arabidopsis leaves. This suggests that the induction of ABA‐related gene expression could be an important component of the Arabidopsis–aphid interaction. Consistent with this hypothesis, M. persicae populations induced ABA production in wild‐type plants. Furthermore, aphid populations were smaller on Arabidopsis aba1‐1 mutants, which cannot synthesize ABA, and showed a significant preference for wild‐type plants compared with the mutant. Total free amino acids, which play an important role in aphid nutrition, were not altered in the aba1‐1 mutant line, but the levels of isoleucine (Ile) and tryptophan (Trp) were differentially affected by aphids in wild‐type and mutant plants. Recently, indole glucosinolates have been shown to promote aphid resistance in Arabidopsis. In this study, 4‐methoxyindol‐3‐ylmethylglucosinolate was more abundant in the aba1‐1 mutant than in wild‐type Arabidopsis, suggesting that the induction of ABA signals that decrease the accumulation of defence compounds may be beneficial for aphids.     

Widespread Shortening of 3’ Untranslated Regions and Increased Exon Inclusion Are Evolutionarily Conserved Features of Innate Immune Responses to Infection

The contribution of pre-mRNA processing mechanisms to the regulation of immune responses remains poorly studied despite emerging examples of their role as regulators of immune defenses. We sought to investigate the role of mRNA processing in the cellular responses of human macrophages to live bacterial infections. Here, we used mRNA sequencing to quantify gene expression and isoform abundances in primary macrophages from 60 individuals, before and after infection with Listeria monocytogenes and Salmonella typhimurium. In response to both bacteria we identified thousands of genes that significantly change isoform usage in response to infection, characterized by an overall increase in isoform diversity after infection. In response to both bacteria, we found global shifts towards (i) the inclusion of cassette exons and (ii) shorter 3’ UTRs, with near-universal shifts towards usage of more upstream polyadenylation sites. Using complementary data collected in non-human primates, we show that these features are evolutionarily conserved among primates. Following infection, we identify candidate RNA processing factors whose expression is associated with individual-specific variation in isoform abundance. Finally, by profiling microRNA levels, we show that 3’ UTRs with reduced abundance after infection are significantly enriched for target sites for particular miRNAs. These results suggest that the pervasive usage of shorter 3’ UTRs is a mechanism for particular genes to evade repression by immune-activated miRNAs. Collectively, our results suggest that dynamic changes in RNA processing may play key roles in the regulation of innate immune responses.     

Epistatic Gene-Based Interaction Analyses for Glaucoma in eMERGE and NEIGHBOR Consortium

Primary open angle glaucoma (POAG) is a complex disease and is one of the major leading causes of blindness worldwide. Genome-wide association studies have successfully identified several common variants associated with glaucoma; however, most of these variants only explain a small proportion of the genetic risk. Apart from the standard approach to identify main effects of variants across the genome, it is believed that gene-gene interactions can help elucidate part of the missing heritability by allowing for the test of interactions between genetic variants to mimic the complex nature of biology. To explain the etiology of glaucoma, we first performed a genome-wide association study (GWAS) on glaucoma case-control samples obtained from electronic medical records (EMR) to establish the utility of EMR data in detecting non-spurious and relevant associations; this analysis was aimed at confirming already known associations with glaucoma and validating the EMR derived glaucoma phenotype. Our findings from GWAS suggest consistent evidence of several known associations in POAG. We then performed an interaction analysis for variants found to be marginally associated with glaucoma (SNPs with main effect p-value <0.01) and observed interesting findings in the electronic MEdical Records and GEnomics Network (eMERGE) network dataset. Genes from the top epistatic interactions from eMERGE data (Likelihood Ratio Test i.e. LRT p-value <1e-05) were then tested for replication in the NEIGHBOR consortium dataset. To replicate our findings, we performed a gene-based SNP-SNP interaction analysis in NEIGHBOR and observed significant gene-gene interactions (p-value <0.001) among the top 17 gene-gene models identified in the discovery phase. Variants from gene-gene interaction analysis that we found to be associated with POAG explain 3.5% of additional genetic variance in eMERGE dataset above what is explained by the SNPs in genes that are replicated from previous GWAS studies (which was only 2.1% variance explained in eMERGE dataset); in the NEIGHBOR dataset, adding replicated SNPs from gene-gene interaction analysis explain 3.4% of total variance whereas GWAS SNPs alone explain only 2.8% of variance. Exploring gene-gene interactions may provide additional insights into many complex traits when explored in properly designed and powered association studies.     

Senescence‐associated tissue microenvironment promotes colon cancer formation through the secretory factor GDF15

The risk of colorectal cancer (CRC) varies between people, and the cellular mechanisms mediating the differences in risk are largely unknown. Senescence has been implicated as a causative cellular mechanism for many diseases, including cancer, and may affect the risk for CRC. Senescent fibroblasts that accumulate in tissues secondary to aging and oxidative stress have been shown to promote cancer formation via a senescence‐associated secretory phenotype (SASP). In this study, we assessed the role of senescence and the SASP in CRC formation. Using primary human colon tissue, we found an accumulation of senescent fibroblasts in normal tissues from individuals with advanced adenomas or carcinomas in comparison with individuals with no polyps or CRC. In in vitro and ex vivo model systems, we induced senescence using oxidative stress in colon fibroblasts and demonstrated that the senescent fibroblasts secrete GDF15 as an essential SASP factor that promotes cell proliferation, migration, and invasion in colon adenoma and CRC cell lines as well as primary colon organoids via the MAPK and PI3K signaling pathways. In addition, we observed increased mRNA expression of GDF15 in primary normal colon tissue from people at increased risk for CRC in comparison with average risk individuals. These findings implicate the importance of a senescence‐associated tissue microenvironment and the secretory factor GDF15 in promoting CRC formation.     

Plasma proteomics and the paediatric patient

Introduction: Plasma proteomics has been extensively utilized for studies that investigate various disease settings (e.g. cardiovascular disease), as well as to monitor the effect of pharmaceuticals on the plasma proteome (e.g. chemotherapy). However, plasma proteomic studies focusing on children represent a very small proportion of the plasma proteomic studies completed to date. Early disease detection and prevention is critical in pediatrics, as children must live with the disease outcomes for many years and often carry negative outcomes into adulthood. Pediatrics represents an area of plasma proteomics that is about to undergo a significant expansion.

Areas covered: This review is based on a PubMed search focusing on five keywords that are plasma, biomarkers, pediatric, proteomics, and children. It is a comprehensive summary of plasma proteomic studies specific to the pediatric patient and discusses aspects such as the clinical setting, sample size, methodological approaches and outlines the significance of the findings.

Expert commentary: Plasma proteomics is expanding significantly as a result of major advancements in proteomic technology. This is in synergy with the growing focus on true early disease detection and prevention in early life. We are about to see a new era of advanced medical science built from pediatric proteomics.