Identification, quantification, and evolutionary analysis of a novel isoform of MCM9

The minichromosome maintenance (MCM) family of proteins is conserved from archaea to humans and is required for assembly of pre-replication complexes (pre-RCs) to initiate DNA replication. MCM9 is an uncharacterized member of the eukaryotic MCM protein family that contains conserved ATP binding and hydrolysis motifs. We have identified a novel alternatively spliced isoform of HsMCM9 that results in a medium length protein product (MCM9M) that eliminates a long C-terminal extension of the fully spliced product (MCM9L). Quantitative real-time reverse transcriptase PCR (qRT-PCR) separated and measured the relative mRNA isoform expression levels across a variety of cell lines. Although there is some variability in expression levels, the full length MCM9L transcript is more abundant than the MCM9M variant in all cell lines tested. The expression of both MCM9 isoforms is cell cycle regulated: induced in S-phase, decreases through G2/M, and becomes constant through G1. Consistent with recent reports suggesting MCM9 participates in repair or prevention of double strand breaks, mitomycin C significantly induces the specific expression of MCM9L, while the replication fork inhibitor, hydroxyurea, has no effect. Evolutionary analysis indicates that the MCM9M isoform is a conserved variant, whereas the addition of the terminal exon producing MCM9L appears to be a more recent event present only in the highest order of eukaryotes.