All known orthologs of a secondary wall-associated cellulose synthase (CesA) gene from Arabidopsis, AtCesA8, encode CesA proteins with two consecutive methionines at their N-termini (MM or 2M). Here, we report that these 2Ms in an aspen ortholog of AtCesA8, PtdCesA8A, are important for maintaining normal wood cellulose biosynthesis in aspen trees. Overexpression of an altered PtdCesA8A cDNA encoding a PtdCesA8A protein missing one methionine at the N-terminus (1M) in aspen resulted in substantial decrease in cellulose content and caused negative effects on wood strength, suggesting that both methionines are essential for proper CesA expression and function in developing xylem tissues. Transcripts from a pair of paralogous native PtdCesA8 genes, as well as introduced PtdCesA8A:1M transgenes were significantly reduced in developing xylem tissues of transgenic aspen plants, suggestive of a co-suppression event. Overexpression of a native PtdCesA8A cDNA encoding a CesA protein with 2Ms at the N-terminus did not cause any such phenotypic changes. These results suggest the importance of 2Ms present at the N-terminus of PtdCesA8A protein during cellulose synthesis in aspen.