Establishment of Repertoire of Placentome-Associated MicroRNAs and Their Appearance in Blood Plasma Could Identify Early Establishment of Pregnancy in Buffalo (Bubalus bubalis).

Precise early pregnancy diagnosis in dairy animals is of utmost importance for an efficient dairy production system. Not detecting a dairy animal pregnant sufficiently early after the breeding results to extending the unproductive time of their milk production cycle and causes substantial economic loss for a dairy producer. At present, the most conventional and authentic pregnancy confirmation practice in cows and buffaloes is rectal palpation of the reproductive organs at Days 35-40 after insemination, which sometime leads to considering an animal as false pregnant. Other alternative methods available for early pregnancy diagnosis lack either accuracy or reproducibility or require elaborate instrumentation and laboratory setup not feasible to practice at farmers' doorstep. The present study was aimed at establishment of the microRNA (miRNA) repertoire of the placentome in buffaloes, which could capture the event of the cross talk between a growing embryo and a dam, through fetal cotyledons and maternal caruncles, and thus could hint at the early pregnancy establishment event in ruminants. Total RNA was isolated from buffalo placentome tissues during early stages of pregnancy (at Day < 25 and Days 30-35), and global small RNA analysis was performed by using Illumina single-end read chemistry and Bubalus bubalis genome. A total of 2,199 miRNAs comprising 1,620 conserved and 579 non-conserved miRNAs were identified. Stringent functional miRNA selection criteria could predict 20 miRNAs worth evaluating for their abundance in the plasma of pregnant, non-pregnant, cyclic non-bred, and non-cyclic prepubertal animals. Eight of them (viz., miR-195-5p, miR-708-3p, miR-379-5p, miR-XX1, miR-XX2, miR-130a-3p, miR-200a-3p, and miR-27) displayed typical abundance patterns in the plasma samples of the animals on Day 19 as well as Day 25 post-insemination, thus making them ambiguous candidates for early pregnancy detection. Similarly, higher abundance of miR-200a-3p and miR130a-3p in non-pregnant animals was indicative of their utility for detecting the animals as not pregnant. Most interestingly, miR-XX1 and miR-XX2 were very characteristically abundant only in pregnant animals. In silico target prediction analysis confirmed that these two miRNAs are important regulators of cyclooxygenase-2 (COX-2) and cell adhesion molecule-2 (CADM-2), both of which play a significant role in the implantation process during feto-maternal cross talk. We interpret that circulatory miR-XX1 and miR-XX2 in blood plasma could be the potential biomarkers for early pregnancy detection in buffaloes.

Unusual interplay of contrasting selective pressures on ß-defensin genes implicated in male fertility of the Buffalo (Bubalus bubalis).

BACKGROUND: The buffalo, despite its superior milk-producing ability, suffers from reproductive limitations that constrain its lifetime productivity. Male sub-fertility, manifested as low conception rates (CRs), is a major concern in buffaloes. The epididymal sperm surface-binding proteins which participate in the sperm surface remodelling (SSR) events affect the survival and performance of the spermatozoa in the female reproductive tract (FRT). A mutation in an epididymal secreted protein, beta-defensin 126 (DEFB-126/BD-126), a class-A beta-defensin (CA-BD), resulted in decreased CRs in human cohorts across the globe. To better understand the role of CA-BDs in buffalo reproduction, this study aimed to identify the BD genes for characterization of the selection pressure(s) acting on them, and to identify the most abundant CA-BD transcript in the buffalo male reproductive tract (MRT) for predicting its reproductive functional significance. RESULTS: Despite the low protein sequence homology with their orthologs, the CA-BDs have maintained the molecular framework and the structural core vital to their biological functions. Their coding-sequences in ruminants revealed evidence of pervasive purifying and episodic diversifying selection pressures. The buffalo CA-BD genes were expressed in the major reproductive and non-reproductive tissues exhibiting spatial variations. The Buffalo BD-129 (BuBD-129) was the most abundant and the longest CA-BD in the distal-MRT segments and was predicted to be heavily O-glycosylated. CONCLUSIONS: The maintenance of the structural core, despite the sequence divergence, indicated the conservation of the molecular functions of the CA-BDs. The expression of the buffalo CA-BDs in both the distal-MRT segments and non-reproductive tissues indicate the retention the primordial microbicidal activity, which was also predicted by in silico sequence analyses. However, the observed spatial variations in their expression across the MRT hint at their region-specific roles. Their comparison across mammalian species revealed a pattern in which the various CA-BDs appeared to follow dissimilar evolutionary paths. This pattern appears to maintain only the highly efficacious CA-BD alleles and diversify their functional repertoire in the ruminants. Our preliminary results and analyses indicated that BuBD-129 could be the functional ortholog of the primate DEFB-126. Further studies are warranted to assess its molecular functions to elucidate its role in immunity, reproduction and fertility.

Integrated analysis of dysregulated lncRNA expression in breast cancer cell identified by RNA-seq study.

Among all the sequencing techniques, RNA sequencing (RNA-seq) has galloped with pace adopting the profiling of transcriptomic data in almost every biological analytics area like gene regulation study, development biology and clinical research. Recently the discovery of differentially expressed genes across different conditions has outshone the barrier of genetic & epigenetic regulations. The present work identified and analyzed differentially expressed novel long non-coding RNAs (lncRNAs) for breast cancer. A complex computational pipeline was adopted for the study which includes analysis of 18498 differentially expressed genes with 4114 up-regulated and 3475 down-regulated transcripts. The overexpression of lnc-MTAP (CDKN2B-AS1), lnc-PCP4 (DSCAM-S1), and lnc-FAM (H19) in breast cells suggests that these lncRNAs may have significant role to play in breast cancer. These results validated the relevance of the dysregulation pattern in cancer cells due to the presence of lncRNAs. The study further opens a new scope for experimental analysis to confirm the aberrant expression pattern of these lncRNAs which may act as potential bio-markers for the diagnosis and early detection of breast cancer.