Possible role of calcium dependent protein phosphorylation in the modulation of wound induced HRGP gene activation in potatoes after gamma irradiation.

Hydroxyproline rich glycoprotein (HRGP) gene is induced in both control and gamma irradiated potato tubers after wounding. The enhanced RNA synthesis in response to wounding correlated well with the accumulation of both HRGP gene transcripts and protein. Initially, the level of HRGP gene expression in gamma irradiated potatoes in response to wounding was 30% more than the corresponding controls. After post irradiation storage of 3-5 weeks, HRGP gene expression in response to wounding was significantly lower than the unirradiated samples. This low level of HRGP gene expression in irradiated potatoes was partially retrieved by 5 mM Ca2+ treatment. Prior treatment with trifluoperazine, a calcium channel blocker resulted in 35% reduction in wound induced HRGP gene expression in control potatoes, further providing evidence for the involvement of Ca2+ dependency for HRGP gene activation. A comparative study on in vivo protein phosphorylation induced by wounding in control and irradiated potatoes exhibited significant differences. A good correlation was observed in the modulation of phosphorylation and HRGP gene expression by Ca2+ in irradiated potatoes. Wound induced signal transduction system and subsequent Ca2+ dependent protein phosphorylation for the activation of HRGP gene is affected in potatoes after gamma irradiation, thus impairing the wound healing process adversely.

Newly synthesised mRNA as a probe for identification of wound responsive genes from potatoes.

A method for the selective isolation of wound induced RNA from potato tuber has been standardised by labeling with 4-thiouridine and subsequent affinity chromatography on Affigel 501 phenyl-mercury Sepharose column. About 30% of the total RNA isolated from wounded potato tissue was found to be synthesized de novo. The relative abundance of wound induced RNAs was confirmed by using two well identified wound inducible gene probes of hydroxyproline-rich glycoproteins (HRGP) and proteinase inhibitor I. The selectively separated wound induced mRNA was used to make cDNA probes. By screening a--gt10 cDNA library from wounded potato tissue with these cDNA probes, several wound responsive genes were identified. Wound inducible expression of these genes was further confirmed by Northern hybridization and their tissue specific expressions were examined. Thus, a simple method for the direct identification of wound responsive genes from potato was developed.