Molecular characterization and post-transcriptional regulation of ME1, a type-I ribosome-inactivating protein from Mirabilis expansa.
Ribosome-inactivating proteins (RIPs) are N-glycosidases that remove a specific adenine from the sarcin/ricin (S/R) loop of the large rRNA, thus arresting protein synthesis at the translocation step. In the present study, ME1, a type-1 RIP, was cloned and sequenced from storage roots of Mirabilis expansa (Ruiz & Pavon). The full-length cDNA sequence of ME1 has 1,129 nucleotides with an open reading frame of 951 nucleotides representing 317 amino acids. Nucleotide analysis revealed that the N-terminal region of ME1 was cleaved, and the mature protein started at amino acid 34. ME1 showed very close similarities to MAP and MAP-4 from Mirabilis jalapa. Southern blot analysis revealed the presence of two homologous genes for ME1 cDNA in M. expansa. Northern blot analysis showed high levels of ME1 transcripts in primary and storage roots. Interestingly, jasmonic acid induced ME1 transcript expression in cell suspension cultures of M. expansa; however, the production of ME1 protein was not enhanced as observed by Western blot analysis. Our data suggest that ME1 has the ability to depurinate its own mRNA, thus inhibiting its translation. These observations suggest a possible mechanism by which ME1 protein levels are post-transcriptionally regulated.
Department of Horticulture and Landscape Architecture, Cell and Molecular Biology Graduate Program, Colorado State University, Fort Collins 80523-1173, USA.
Ramarao
R
Vepachedu
Harsh Pal
HP
Bais
Jorge M
JM
Vivanco
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015139:Blotting, Southern; D002478:Cells, Cultured; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics; D018506:Gene Expression Regulation, Plant; D039282:Mirabilis / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D010940:Plant Proteins / Q000235:genetics* / Q000378:metabolism / Q000494:pharmacology; D018517:Plant Roots / Q000235:genetics / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D011817:Rabbits; D012156:Reticulocytes / Q000187:drug effects; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid
D018076:DNA, Complementary; D010940:Plant Proteins; D012333:RNA, Messenger