In this talk, I will try to provide evidence that alternative splicing of pre-mRNA and non-canonical binding of microRNA (miRNA) can regulate the function of MYH genes which encode heavy chains of nonmuscle myosin II (NM II). First half of my talk will discuss a novel alternatively spliced isoform of NM IIA, called NM IIA2, which is generated by the inclusion of 21 amino acids near the actin binding region (loop 2) of the head domain of heavy chain which is encoded by MYH9 gene. Expression of NM IIA2 is found exclusively in the brain tissue, where it reaches a maximum level at 24h during the circadian rhythm. The actin-dependent Mg2+-ATPase activity and in vitro motility assays reveal that NM IIA2 lacks its motor activities but localizes with actin filaments in cells. Interestingly, NM IIA2 can also make heterofilaments with NM IIA0 (non-inserted isoform of NM IIA) and can retard the in vitro motility of NM IIA, when the two are mixed, suggesting the functional importance of alternatively spliced isoform, NM IIA2, and its potential physiological role in regulating NM IIA activity in the brain. Second half of my talk will discuss that a group of microRNAs (mmu-miR-200a-5p, mmu-miR-532-3p, mmu-miR-680 and mmu-miR-1901) can significantly repress the expression of NMIIC (a paralog of NMIIA). Interestingly, these microRNAs have both canonical and non-canonical binding sites at 3/UTR and CDS of NMIIC’s heavy chain (HC) mRNA, encoded by MYH14 gene. Each of the miRNA downregulates NMHC-IIC to a different degree as assessed by dual-luciferase and immunoblot analyses. When we abolish the complementary base pairing at canonical binding site, mmu-miR-532-3p can still bind at non-canonical binding site and form Argonaute 2 (AGO2)-miRNA complex to downregulate the expression of NMIIC. Together, these studies suggest that MYH genes’ activity can be regulated by their alternative splicing and abundance of miRNA targeting them.