50 Years of Image Analysis

The COVID-19 pandemic is an unprecedented global challenge, and point-of-care diagnostic classifiers are urgently required. Here, we present a platform for ultra-high-throughput serum and plasma proteomics that builds on ISO13485 standardization to facilitate simple implementation in regulated clinical laboratories. Our low-cost workflow handles up to 180 samples per day, enables high precision quantification, and reduces batch effects for large-scale and longitudinal studies. We use our platform on samples collected from a cohort of early hospitalized cases of the SARS-CoV-2 pandemic and identify 27 potential biomarkers that are differentially expressed depending on the WHO severity grade of COVID-19. They include complement factors, the coagulation system, inflammation modulators, and pro-inflammatory factors upstream and downstream of interleukin 6. All protocols and software for implementing our approach are freely available. In total, this work supports the development of routine proteomic assays to aid clinical decision making and generate hypotheses about potential COVID-19 therapeutic targets.

Ultra-High-Throughput Clinical Proteomics Reveals Classifiers of COVID-19 Infection.

The serpin family comprises a structurally similar, yet functionally diverse, set of proteins. Named originally for their function as serine proteinase inhibitors, many of its members are not inhibitors but rather chaperones, involved in storage, transport, and other roles. Serpins are found in genomes of all kingdoms, with 36 human protein-coding genes and five pseudogenes. The mouse has 60 Serpin functional genes, many of which are orthologous to human SERPIN genes and some of which have expanded into multiple paralogous genes. Serpins are found in tissues throughout the body; whereas most are extracellular, there is a class of intracellular serpins. Serpins appear to have roles in inflammation, immune function, tumorigenesis, blood clotting, dementia, and cancer metastasis. Further characterization of these proteins will likely reveal potential biomarkers and therapeutic targets for disease.

https://link.springer.com/content/pdf/10.1186%2F1479-7364-7-22.pdf

Update of the human and mouse SERPIN gene superfamily.

Chemogenetic characterization through in vitro evolution combined with whole-genome analysis can identify antimalarial drug targets and drug-resistance genes. We performed a genome analysis of 262 Plasmodium falciparum parasites resistant to 37 diverse compounds. We found 159 gene amplifications and 148 nonsynonymous changes in 83 genes associated with drug-resistance acquisition, where gene amplifications contributed to one-third of resistance acquisition events. Beyond confirming previously identified multidrug-resistance mechanisms, we discovered hitherto unrecognized drug target-inhibitor pairs, including thymidylate synthase and a benzoquinazolinone, farnesyltransferase and a pyrimidinedione, and a dipeptidylpeptidase and an arylurea. This exploration of the P. falciparum resistome and druggable genome will likely guide drug discovery and structural biology efforts, while also advancing our understanding of resistance mechanisms available to the malaria parasite.

/pdf/mapping-the-malaria-parasite-druggable-genome-by-using-in-4fxp33wbqp.pdf

Mapping the malaria parasite druggable genome by using in vitro evolution and chemogenomics

Recurrent miscarriage (RM) occurs in 1-3% of couples aiming at childbirth. Due to multifactorial etiology the clinical diagnosis of RM varies. The design of genetic/’omics’ studies to identify genes and biological mechanisms involved in pathogenesis of RM has challenges as there are several options in defining the study subjects (female patient and/or couple with miscarriages, fetus/placenta) and controls. An ideal study would attempt a trio-design focusing on both partners as well as pregnancies of the couple. Application of genetic association studies focusing on pre-selected candidate genes with potential pathological effect in RM show limitations. Polymorphisms in ~100 genes have been investigated and association with RM is often inconclusive or negative. Also, implication of prognostic molecular diagnostic tests in clinical practice exhibits uncertainties. Future directions in investigating biomolecular risk factors for RM rely on integrating alternative approaches (SNPs, CNVs, gene/protein expression, epigenetic regulation) in studies of single genes as well as whole-genome analysis. This would be enhanced by collaborative network between research centers and RM clinics.

/pdf/genetics-of-recurrent-miscarriage-challenges-current-1iz3isdm69.pdf

Genetics of recurrent miscarriage: challenges, current knowledge, future directions.

Objective. Vascular endothelial growth factor (VEGF) is a pro-angiogenic factor. Variability in VEGF expression, induced by specific VEGFA variants, are involved in angiogenesis-related disorders. This study examined the genotype distribution and functional role (VEGF expression) of rs699947, rs833061, rs1570360, rs2010963, rs833068, rs833070, rs3025020, and rs3025039 VEGFA variants and their haplotypes in 519 healthy Bahraini individuals of both genders. Methods and results. The distribution of the eight VEGFA polymorphisms screened was in Hardy-Weinberg equilibrium. The minor allele frequencies of rs699947 (0.42), rs833061 (0.32), rs1570360 (0.31), rs2010963 (0.33), rs833068 (0.37), rs833070 (0.42), rs3025020 (0.33), and rs3025039 (0.13) were generally compared to those established for Caucasians. Of the variants tested, rs3025020 was associated with increased VEGF serum levels (p=0.019), while rs3025039 was associated with decreased levels (p=0.038). Linkage analysis identified two VEGFA blocks, the first, spanning 16 kb, was not associated with altered VEGF levels, while the second, spanning 3 kb containing rs3025020 and rs3025039, was linked with higher VEGF expression, of which the -583 T / +936 T haplotype (p=0.008) was linked with higher VEGF levels compared to the -583C/ +936C (all wild-type) haplotype. Conclusion. These results support the association of rs30250202 and rs3025039, and specific VEGF haplotypes, with altered VEGF serum levels, although the exact functional mechanisms remain to be elucidated.

Contribution of VEGF polymorphisms to variation in VEGF serum levels in a healthy population

Forkhead-associated (FHA) domains are small phosphopeptide recognition modules found in eubacterial and eukaryotic, but not archeal, genomes. Although they were originally found in forkhead-type transcription factors, they have now been identified in many other signaling proteins. FHA domains share a remarkably conserved fold despite very low sequence conservation. They only have five conserved amino acids that are important for binding to phosphorylated epitopes. Recent work from several laboratories has demonstrated that FHA domains can mediate many interactions that do not depend on their ability to recognize a phosphorylated threonine. In this review, we present structural and biochemical work that has unveiled novel interaction interfaces on FHA domains. We discuss how these non-canonical interactions modulate the recognition of phosphorylated and non-phosphorylated substrates, as well as protein oligomerization - events that collectively determine FHA function.

FHA domains: Phosphopeptide binding and beyond

In conditions of proteasomal impairment, the build-up of damaged or misfolded proteins activates a cellular response leading to the recruitment of damaged proteins into perinuclear aggregates called aggresomes. Aggresome formation involves the retrograde transport of cargo proteins along the microtubule network and is dependent on the histone deacetylase HDAC6. Here we show that ionizing radiation (IR) promotes Ran-Binding Protein M (RanBPM) relocalization into discrete perinuclear foci where it co-localizes with aggresome components ubiquitin, dynein and HDAC6, suggesting that the RanBPM perinuclear clusters correspond to aggresomes. RanBPM was also recruited to aggresomes following treatment with the proteasome inhibitor MG132 and the DNA-damaging agent etoposide. Strikingly, aggresome formation by HDAC6 was markedly impaired in RanBPM shRNA cells, but was restored by re-expression of RanBPM. RanBPM was found to interact with HDAC6 and to inhibit its deacetylase activity. This interaction was abrogated by a RanBPM deletion of its LisH/CTLH domain, which also prevented aggresome formation, suggesting that RanBPM promotes aggresome formation through an association with HDAC6. Our results suggest that RanBPM regulates HDAC6 activity and is a central regulator of aggresome formation.

/pdf/aggresome-formation-is-regulated-by-ranbpm-through-an-6b8dst2x7a.pdf

Ahmad W. Almawi

Papers

Contribution of VEGF polymorphisms to variation in VEGF serum levels in a healthy population

FHA domains: Phosphopeptide binding and beyond

Aggresome formation is regulated by RanBPM through an interaction with HDAC6.

Replication study of common variants in CDKAL1 and CDKN2A/2B genes associated with type 2 diabetes in Lebanese Arab population

Strong association of common variants in the IGF2BP2 gene with type 2 diabetes in Lebanese Arabs