Systems biology approach to study the role of miRNA in promoter targeting during megakaryopoiesis

Highlights: • miRNAs could regulate Cyclins and CDKs by targeting their promoters. • Perl programming was used to identify putative targets of miRNA in promoters. • miR-1273g-3p and miR-619-5p showed significance with longest seed sequence matches. • Through CDK10, CDK11 and Cyclin, these two miRNAs could regulate cell cycle during megakaryopoiesis. The distinct process of megakaryopoiesis requires occurrence of endomitosis for polyploidization of the megakaryocytes. Although, Cyclins, CDKs and have been described to regulate endomitosis, the exact mechanism still remains an enigma. miRNA which were otherwise known as post transcriptional gene silencers are now emerging with various non-canonical functions including gene regulation at pre-transcriptional level by miRNA binding at promoter region. Out of the many processes they regulate, miRNA have been manifested to play a role in megakaryocyte differentiation. In this study an attempt has been made to identify miRNA that could regulate cell cycle genes (Cyclins and CDKs) by targeting their promoters, during megakaryopoiesis. A new computational algorithm was implemented using Perl programming to identify putative targets of miRNA in CDK and Cyclin promoters. Perl script was also used to check nuclear localizing miRNA based on the presence of a consensus sequence. Real-time PCR was performed to analyze the expression of miRNA and their predicted targets in Dami vs. PMA treated Dami cells. Putative targets of miRNAs with longest, high complementarity matches in CDK/Cyclin promoters were obtained. We identified two significant miRNA, miR-1273g-3p and miR-619-5p with longest seed sequence matches. We further identified three main targets (CDK10, CDK11, Cyclin F) through which these two miRNA could regulate cell cycle during megakaryopoiesis. Our results reinforce the role of promoting targeting miRNA in regulation of cell cycle through certain CDK/Cyclins to support the process of endomitosis during megakaryopoiesis.