Expression profiling and in silico characterization of bubaline Drosha in light of evolution

Drosha is a class III ribonuclease and is a critical enzyme involved in miRNA biogenesis. The rationale behind the current study was to profile the tissue specific expression of bubaline Drosha and envisage its evolution in the light of other RNases of animal and prokaryotic origin. Relative quantification using qRT-PCR (TaqMan chemistry) revealed that its expression was the highest in kidney and heart and the lowest in brain. The clones of the overlapping partial cds of bubaline Drosha were custom sequenced. The sequences of Drosha and Dicer transcript variants of divergent origin were subjected to biocomputational analyses (using MEGA6 and Datamonkey server) which indicated that the bovidae, suidae, and marine mammals form different phylogenetic clusters. Wide range of variation among the divergent species indicated natural selection to confer specific functionality. To identify the selection in codons of the Drosha and Dicer the Ribonuclease domains of representative divergent sequences were analyzed using different models (SLAC, REL, FEL) of Datamonkey server. The salient findings were that certain codons of RNases have undergone purifying selection during evolution, the type III RNases have evolved independently of the RNase-L, -A and –H of animal origin from respective progenitors in the prokaryotic RNases and the domains of different RNases different markedly from each other as evidenced by the protein structure analyses. Finally, the divergent RNases were subjected functional classification and network analyses (Panther and String-db online tools) to identify their inter-relationship with respect to their functions. It was evident that the RNases of prokaryotic vis-a-vis animal origin are involved in specific networks according to the functional similarity. The overall findings indicate that the prokaryotic RNases have evolved with respect to the functional requirement and has acted as the progenitor of the more divergent animal RNases.