Developmental and light-regulated post-translational control of 3-hydroxy-3-methylglutaryl-CoA reductase levels in potato.
In plants, the first committed step in the cytosolic pathway for biosynthesis of isoprenoids is catalysed by 3-hydroxy-3-methylglutaryl-CoA reductase (HMGR). We have added an eight amino-acid-residue epitope tag to a potato (Solanum tuberosum L.) HMGR isoform and expressed this novel protein (HMGR-FLAG) in transgenic plants. Despite high levels of transcript accumulation in all leaf stages of transgenic plants, high levels of HMGR-FLAG protein were found only in apical meristematic tissue, suggesting post-translational regulation of potato HMGR affected by plant development. Protein immunoblots, and determination of enzymatic activity and transcript accumulation in the HMGR-FLAG transgenic and the non-transgenic parental plant lines, show that HMGR levels decrease dramatically in the dark. Again, the mechanism of this control occurs at a post-translational level. After 2.5 h in darkness, levels of HMGR-FLAG are approximately one-half of those in plants in the light; protein levels recover rapidly when dark-treated plants are returned to the light. In non-transgenic plants, hmg transcript levels are reduced in the dark, whereas dark treatments do not affect transgene hmg transcripts expressed under the control of a constitutive promoter. Furthermore, transcripts for HMGR-FLAG remain associated with polyribosomes in dark-treated tissues. Addition of inhibitors of cysteine proteases during microsomal protein extraction is required for recovery of immunoreactive HMGR-FLAG. The epitope-tagged isozyme has been used to show for the first time that a regulated decrease in plant HMGR activity correlates closely with a loss of the HMGR protein. We have used whole plants to demonstrate that developmental and light-regulated control of HMGR occurs post-translationally in vivo.
Korth
K L
KL
Department of Plant Pathology, 217 Plant Science Building, University of Arkansas, Fayetteville, AR 72701, USA. kkorth@comp.uark.edu
Jaggard
D A
DA
Dixon
R A
RA
D015152:Blotting, Northern; D003624:Darkness; D018507:Gene Expression Regulation, Developmental; D018506:Gene Expression Regulation, Plant; D006903:Hydroxymethylglutaryl CoA Reductases / Q000235:genetics* / Q000378:metabolism; D015151:Immunoblotting; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism; D008027:Light; D018515:Plant Leaves / Q000201:enzymology / Q000235:genetics / Q000254:growth & development; D030821:Plants, Genetically Modified; D014176:Protein Biosynthesis; D011198:Solanum tuberosum / Q000201:enzymology / Q000235:genetics* / Q000254:growth & development
D007527:Isoenzymes; D006903:Hydroxymethylglutaryl CoA Reductases