Male Sterile2 Encodes a Plastid-Localized Fatty Acyl Carrier Protein Reductase Required for Pollen Exine Development in Arabidopsis

Male Sterile 2 (MS2) is predicted to encode a fatty acid reductase required for pollen wall development in Arabidopsis thaliana. Transient expression of MS2 in tobacco leaves resulted in the accumulation of significant levels of C16 and C18 fatty alcohols. Expression of MS2 fused with GFP revealed that an N-terminal transit peptide targets the MS2 to plastids. The plastidial localization of MS2 is biologically important because genetic complementation of MS2 in ms2 homozygous plants was dependent on the presence of its N-terminal transit peptide or that of Rubisco small subunit protein N-terminal transit peptide (RTP). In addition, two domains: (NAD(P)H binding domain) (NBD) and sterile domain (SD), conserved in MS2 and its homologs, were also shown to be essential for MS2 function in pollen exine development by genetic complementation testing. Direct biochemical analysis revealed that purified recombinant MS2 enzyme is able to convert palmiltoyl-ACP to the corresponding C16:0 alcohol with NAD(P)H as the preferred electron donor. Using optimized reaction conditions i.e., at pH6.0 and 30°C, MS2 exhibits a Km for 16:0-ACP of 23.3±4.0 µM, a Vmax of 38.3±4.5 nmol mg-1 min-1 and a kcat /Km of 1,873 M-1s-1. Based on the high homology of MS2 to other characterized fatty acid reductases, it was surprising that MS2 showed no activity against palmitoyl- or other acyl-CoAs, this is however consistent with its plastidial localization. In summary, genetic and biochemical evidence demonstrate an MS2-mediated conserved plastidial pathway for the production of fatty alcohols that are essential for pollen wall biosynthesis in Arabidopsis.