Isolation and characterization of a phospholipase C delta isoform from pea that is regulated by light in a tissue specific manner.
Phosphoinositide-specific phospholipases C (PLCs) play an important role in many cellular responses and are involved in the production of secondary messengers. We report the cloning and characterization of a cDNA encoding a PLC-delta from Pisum sativum (PsPLC). The amino acid sequence deduced from the cDNA sequence showed 75-80% identity to other plant PLCs and contained the characteristic X, Y and C2 domains. The genomic PLC clone from pea was also characterized and found to contain eight introns. The protein was expressed in Escherichia coli, but the recombinant product did not show any phosphoinositide (PI)- or phosphatidylinositol-4, 5-bisphosphate (PIP2)-specific activity, despite having all known residues required for such activity, and in spite of the fact that its C2 domain was shown to bind calcium. Under similar in vitro assay conditions the recombinant tobacco PLC used as a control showed calcium-dependent PI- and PIP2-specific activity. Though PsPLC did not show enzyme activity in vitro, and may represent an inactive form of PLC, such as those reported in some mammalian systems, analysis of the transcription of PsPLC showed that the gene is expressed in all pea tissues, and is regulated by light in a tissue-specific manner. Roots showed higher expression of PsPLC than shoots. A putative PsPLC promoter region (792 bp) was also cloned and found to contain root-specific and light-responsive cis elements, suggesting that this form of PLC may be involved in important functions in plants.
Venkataraman
G
G
International Centre for Genetic Engineering and Biotechnology (ICEGEB), Aruna Asaf Ali Marg, 110 067 New Delhi, India.
Goswami
M
M
Tuteja
N
N
Reddy
M K
MK
Sopory
S K
SK
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D017931:DNA Primers; D018076:DNA, Complementary; D015971:Gene Expression Regulation, Enzymologic / Q000528:radiation effects*; D018506:Gene Expression Regulation, Plant / Q000528:radiation effects*; D018745:Genome, Plant; D007527:Isoenzymes / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D008027:Light; D008969:Molecular Sequence Data; D018532:Pisum sativum / Q000201:enzymology*; D054803:Phospholipase C delta; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D010738:Type C Phospholipases / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism
D017931:DNA Primers; D018076:DNA, Complementary; D007527:Isoenzymes; D011994:Recombinant Proteins; D010738:Type C Phospholipases; D054803:Phospholipase C delta