Modulation of poly(A) + RNA levels in fungal-infected wheat embryos through the selective inactivation of RNA polymerase II and poly(A) polymerase

Abstract

Infection of germinating wheat embryos by a fungal pathogen ( Drechslera sorokiana) drastically lowered (70–73%) the relative abundance of poly(A) + RNA. This was paralleled by a significant loss in the activities of RNA polymerase II (60–70%) and poly(A) polymerase (80–85%) enzymes. The inhibition of RNA polymerase II (60–65%) and poly(A) polymerase (70–85%) activities was also witnessed by the in vitro addition of the fungal extract to the enzyme preparations isolated from healthy embryos. The fungal extract showed negligible phosphatase and nuclease activities. This ruled out the possibility of rapid degradation of the labelled substrate [ 3H]ATP, primer RNA, or even the labelled reaction products under our assay conditions. The inhibitory effect of the fungal extract could be alleviated by fractionating the treated enzyme preparation by phosphocellulose chromatography. This indicated that the fungal extract was directly responsible for the inactivation of the polymerases in a reversible manner. The inhibitory function of the fungal extract was destroyed by treatment with pronase, but not with RNAase A and RNAase Ti. Poly(A) ‘tails’ were enzymatically excised from 32P-labelled poly(A) + RNA and fractionated on acrylamide gels for autoradiographic analysis. The lengths of the 32P-labelled poly(A) ‘tails’ in control and infected embryos turned out to be identical (64 nucleotides). Our results suggest that the relative abundance of poly(A) + RNA is diminished in fungal-infected wheat embryos through the selective inactivation of RNA polymerase II and poly(A) polymerase enzymes.