To investigate the lignifications process and its physiological significance under Karnal Bunt (KB), the changes in enzymes responsible for lignifications likes, phenylalanine ammonia lyase (PAL), were determined in resistant (HD-29) and susceptible genotype (WH-542) of wheat during different developmental stages. The PAL gene was cloned and sequenced. The expression of PAL gene was measured by means of semi-quantitative RT-PCR. The enzyme was expressed constitutively in both the susceptible and resistant genotype. However, the activity was higher in all the developmental stages of resistant genotype, indicating that this genotype has a significant higher basal level of these enzymes as compared to the susceptible line and could be used as marker(s) to define KB resistance. The activity of PAL was significantly higher in WSv stage (Z=16). Structural comparisons based on alignments of all the protein sequences using the clustal W program and searches for conserved motifs using the MEME program have revealed broad conservation of main motifs characteristic of the plant PAL. MSA and phylogenetic analyses of different plants PAL demonstrate that all PAL cluster divided in to two main cluster. The PAL also possesses a specific consensus sequences [GS]- [STG]-[LIVM]-[STG]-[SAC]-S-G-[DH]-L-x-[PN]-L-[SA]-x(2,3)-[SAGVTL]. The pathway might be associated with the enhancement of structural defense barrier due to lignifications of cell wall as evident from the enhanced synthesis of lignin in all the stages of resistant genotype. Our results clearly indicate the possible role of enzymes of Phenyl propanoid pathway metabolism provides genotype and stage dependant structural barrier resistance in wheat against KB.
Keywords: KB; PAL; Phenyl propanoid pathway; RT PCR; lignifications.