Characterization of the 5 ' region of human CoQ2, a gene causing primary CoQ10 deficiency

Ubiquinone, coenzyme Q(10) or CoQ is a lipid-soluble component of the mitochondrial respiratory chain (RC) where it transfers reducing equivalents from complex I and complex II to complex III. CoQ10biosynthesis is still poor characterized in humans and consists of multiple enzymatic reactions. Primary CoQ10 deficiency causes a from of mitochondrial encephalomyopathy which is inherited as autosomal recessive trait, and in some patients it is associated to mutations in the COQ2 gene, encoding for Para-Hydroxybenzoate-Polyprenyl Transferase (Quinzii et al 2006). Human COQ2 cDNA has been isolated from a human muscle and liver cDNA library (Forsgren et al, 2004) and encodes for a protein located in the mitochondrial inner membrane. The reported human sequence shows four different in frame ATG codons in the 5’ region of the gene. However by aligning the human protein with other mammalian COQ2 proteins, we found that the N-terminus of the reported sequence did not shown any homology with other mammalian COQ2 proteins, moreover there are no GENBANK ESTs corresponding to the reported 5’ region of the gene. We therefore characterized the 5’ region of COQ2 by 5’-RACE and found different transcripts in human fibroblasts, but the most represented one (over 75% of total COQ2 mRNA) included only the fourth ATG. We then amplified cDNA using a fixed primer within exon 2 of the gene, distal to the predicted targeting sequence, and several primers located upstream of the fourth ATG. None of the detected PCR products included the first two reported ATG. To confirm the physiological relevance of these shorter mRNAs, we cloned the PCR fragments in frame with GFP, and we demonstrated that the resulting fluorescent fusion protein is targeted to mitochondria. These data show that the functional human COQ2 mRNA is shorter than what previously was thought. We believe the reported COQ2 transcript represents a very rare mRNA, with little physiological significance. These findings are crucial to perform correct mutation screening in CoQ10 deficient patients.