Synonymous variations, which are defined as codon substitutions that do not change the encoded amino acid, were previously thought to have no effect on the properties of the synthesized protein(s). However, mounting evidence shows that these “silent” variations can have a significant impact on protein expression and function and should no longer be considered “silent”. Here, the effects of six synonymous and six non-synonymous variations, previously found in the gene of ADAMTS13, the von Willebrand Factor (VWF) cleaving hemostatic protease, have been investigated using a variety of approaches. The ADAMTS13 mRNA and protein expression levels, as well as the conformation and activity of the variants have been compared to that of wild-type ADAMTS13. Interestingly, not only the non-synonymous variants but also the synonymous variants have been found to change the protein expression levels, conformation and function. Bioinformatic analysis of ADAMTS13 mRNA structure, amino acid conservation and codon usage allowed us to establish correlations between mRNA stability, RSCU, and intracellular protein expression. This study demonstrates that variants and more specifically, synonymous variants can have a substantial and definite effect on ADAMTS13 function and that bioinformatic analysis may allow development of predictive tools to identify variants that will have significant effects on the encoded protein.

A vast amount of data on the natural resistance of Saccharomyces cerevisiae to a diverse array of chemicals has been generated over the past decade (chemical genetics). We endeavored to use this data to better characterize the “systems” level properties of this phenomenon. By collating data from over 30 different genome-scale studies on growth of gene deletion mutants in presence of diverse chemicals, we assembled the largest currently available gene-chemical network. We also derived a second gene-gene network that links genes with significantly overlapping chemical-genetic profiles. We analyzed properties of these networks and investigated their significance by overlaying various sources of information, such as presence of TATA boxes in their promoters (which typically correlate with transcriptional noise), association with TFIID or SAGA, and propensity to function as phenotypic capacitors. We further combined these networks with ubiquitin and protein kinase-substrate networks to understand chemical tolerance in the context of major post-translational regulatory processes. Hubs in the gene-chemical network (multidrug resistance genes) are notably enriched for phenotypic capacitors (buffers against phenotypic variation), suggesting the generality of these players in buffering mechanistically unrelated deleterious forces impinging on the cell. More strikingly, analysis of the gene-gene network derived from the gene-chemical network uncovered another set of genes that appear to function in providing chemical tolerance in a cooperative manner. These appear to be enriched in lineage-specific and rapidly diverging members that also show a corresponding tendency for SAGA-dependent regulation, evolutionary divergence and noisy expression patterns. This set represents a previously underappreciated component of the chemical response that enables cells to explore alternative survival strategies. Thus, systems robustness and evolvability are simultaneously active as general forces in tolerating environmental variation. We also recover the actual genes involved in the above-discussed network properties and predict the biochemistry of their products. Certain key components of the ubiquitin system (e.g. Rcy1, Wss1 and Ubp16), peroxisome recycling (e.g. Irs4) and phosphorylation cascades (e.g. NPR1, MCK1 and HOG) are major participants and regulators of chemical resistance. We also show that a major subnetwork boosting mitochondrial protein synthesis is important for exploration of alternative survival strategies under chemical stress. Further, we find evidence that cellular exploration of survival strategies under chemical stress and secondary metabolism draw from a common pool of biochemical players (e.g. acetyltransferases and a novel NTN hydrolase).

In the present study, a total of 53 promising salt-tolerant genotypes were tested across 18 salt-affected diverse locations for three years. An attempt was made to identify ideal test locations and mega-environments using GGE biplot analysis. The CSSRI sodic environment was the most discriminating location in individual years as well as over the years and could be used to screen out unstable and salt-sensitive genotypes. Genotypes CSR36, CSR-2K-219, and CSR-2K-262 were found ideal across years. Overall, Genotypes CSR-2K-219, CSR-2K-262, and CSR-2K-242 were found superior and stable among all genotypes with higher mean yields. Different sets of genotypes emerged as winners in saline soils but not in sodic soils; however, Genotype CSR-2K-262 was the only genotype that was best under both saline and alkaline environments over the years. The lack of repeatable associations among locations and repeatable mega-environment groupings indicated the complexity of soil salinity. Hence, a multi-location and multi-year evaluation is indispensable for evaluating the test sites as well as identifying genotypes with consistently specific and wider adaptation to particular agro-climatic zones. The genotypes identified in the present study could be used for commercial cultivation across edaphically challenged areas for sustainable production.

Summary The reproductive‐derived serine protease inhibitor Kazal‐type (Spink) has been identified in seminal plasma, and Spink–spermatozoa binding has been illustrated in many mammalian species including human. We used mice as experimental animal to study the mode of Spink action in the modulation of mammalian sperm activity. A Spink3‐binding zone was cytochemically stained on the sperm head at apical hook separated from intact acrosome, whether the cells were capacitated or not. The Spink3–spermatozoa binding neither changed the population of cells in the uncapacitated, capacitated and acrosome‐reacted status nor affected the capacitation‐related protein phosphorylation and cell motility enhancement. Despite that, the Spink–spermatozoa interaction resulted in decreasing the intracellular calcium concentration ([Ca2+]i) of the cell head and suppressing both the acrosome reaction induced by Ca+2 ionophore A23187 and the cell fertility. Furthermore, Spink3 seen on the head of spermatozoa in the uterine cavity after coitus could be removed by the trypsin‐like activity in the uterine fluid of oestrous females, and free Spink3 in the uterine cavity suppressed the protease activity. We integrated our data to shed light on the molecular mechanism of how Spink and its inhibiting protease are interplayed to modulate the activity of mammalian spermatozoa during their transit in the reproductive tract.

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Germplasm should be conserved in such a way that the genetic integrity of a given accession is maintained. In most genebanks, landraces constitute a major portion of collections, wherein the extent of genetic diversity within and among landraces of crops vary depending on the extent of outcrossing and selection intensity infused by farmers. In this study, we assessed the level of diversity within and among 108 diverse landraces and wild accessions using both phenotypic and genotypic characterization. This included 36 accessions in each of sorghum, pearl millet, and pigeonpea, conserved at ICRISAT genebank. We genotyped about 15 to 25 individuals within each accession, totaling 1,980 individuals using the DArTSeq approach. This resulted in 45,249, 19,052, and 8,211 high-quality single nucleotide polymorphisms (SNPs) in pearl millet, sorghum, and pigeonpea, respectively. Sorghum had the lowest average phenotypic (0.090) and genotypic (0.135) within accession distances, while pearl millet had the highest average phenotypic (0.227) and genotypic (0.245) distances. Pigeonpea had an average of 0.203 phenotypic and 0.168 genotypic within accession distances. Analysis of molecular variance also confirms the lowest variability within accessions of sorghum (26.3%) and the highest of 80.2% in pearl millet, while an intermediate in pigeonpea (57.0%). The effective sample size required to capture maximum variability and to retain rare alleles while regeneration ranged from 47 to 101 for sorghum, 155 to 203 for pearl millet, and 77 to 89 for pigeonpea accessions. This study will support genebank curators, in understanding the dynamics of population within and among accessions, in devising appropriate germplasm conservation strategies, and aid in their utilization for crop improvement.

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