Isolation of pepper mRNAs differentially expressed during the hypersensitive response to tobacco mosaic virus and characterization of a proteinase inhibitor gene

Pepper (Capsicum annuum L. cv. Bugang) displays a hypersensitive response (HR) upon infection by tobacco mosaic virus (TMV) pathotype P₀. To elucidate molecular mechanisms underlying this response, a cDNA library was constructed and differentially screened between TMV-P₀-inoculated and uninoculated leaves. Of 3260 clones analyzed by reverse RNA slot-blot hybridization, 22 showed up-regulated expression upon TMV-P₀ challenge. Among these, 16 clones showed significantly higher levels of expression in response to the avirulent pathotype TMV-P₀ than to the virulent pathotype TMV-P_1.2. Nucleotide sequence analysis and database searches revealed that the polypeptides deduced from the selected clones displayed significant sequence homology to various known proteins, indicating conservation of the wide range of up-regulated proteins during HR against viral challenge in higher plants. A cDNA clone encoding a putative proteinase inhibitor II (CaPinII) was selected and further analyzed. Upon TMV-P₀ inoculation, the CaPinII transcript accumulated maximally at 72 h post-inoculation and declined afterwards. In contrast, inoculation with TMV-P_1.2 did not induce CaPinII expression to any detectable level. The CaPinII gene was also expressed in response to the treatment of various chemicals, as well as wounding. Southern blot analysis showed that a small family of genes homologous to the CaPinII gene is present in the pepper genome.