Identification of multiple genes encoding SnRK1 subunits in potato tuber.

ABSTRACT

BACKGROUND: Many studies have proven the importance of SnRK1 in the regulation of carbohydrate metabolism and plant development. Compared to Arabidopsis, much less is known about SnRK1 complexes in crop plants, and therefore, more work needs to be done to identify SnRK1 genes and to investigate their function in crop plants. METHODS: In this study we identified five SnRK1-related genes in potato by analyzing the potato genome through BLAST, which encode one α-subunit isoform (stKIN), two ß-subunit isoforms (stKINß1 and stKINß2) and two γ-subunit isoforms (stKINγ and stKINßγ). To investigate the functions of SnRK1 in the tuber development of potato, we further made overexpression and RNAi transgenic plants of these five genes. Based on these overexpression transgenic plants, the Fast protein liquid chromatography (FPLC) were employed to purify SnRK1 complexes, which were tracked by western-blot. RESULTS: Experiments in vivo and in vitro showed that these five proteins in potato are functional SNF1/AMPK/SnRK1-related proteins. The SnRK1 activity decreased by 60% in the RNAi transgenic lines of stKIN; the starch content increased by 25% in the overexpression transgenic lines of stKIN, compared to that in the wild-type lines, whereas there is no significant difference in SnRK1 activity and starch content in the RNAi transgenic or overexpression lines of stKINß1, stKINß2, stKINγ and stKINßγ. In addition, we found that a few different SnRK1 complexes are present in potato by partially purifying SnRK1 complexes from these overexpression transgenic plants. CONCLUSIONS: Five functional SnRK1-related genes were identified in potato, including three novel genes, which encode one α-subunit isoform (stKIN), two ß-subunit isoforms (stKINß1 and stKINß2) and two γ-subunit isoforms (stKINγ and stKINßγ). We found that a few SnRK1 related genes are present in potato tuber, which form several different SnRK1 isoenzymes. We found that stKIN is the primary α subunit of SnRK1 in potato tuber and plays important roles in the development of potato tubers.