Expression and functional evaluation of CaZNF830 during pepper response to Ralstonia solanacearum or high temperature and humidity

Extensive transcriptional reprogramming after pathogen attack determines immunity to these invaders and plant development. Zinc finger (ZNF) transcription factors regulate important processes in plants such as development, vegetative activities and plant immunity. Despite their immense significance, majority of ZNF transcription factors (TF) involved in pepper immunity and resistance to heat stress have not been focused much. Herein, we identified and functionally characterized CaZNF830 in pepper defense against Ralstonia solanacearum inoculation (RSI) and tolerance to high temperature and high humidity (HTHH). Transient expression analysis of CaZNF830-GFP fusion protein in tobacco leaves revealed its localization to the nucleus. Transcription of CaZNF830 is induced in pepper plants upon RSI or HTHH. Consistent with this, fluorometric GUS enzymatic assay driven by pCaZNF830 presented significantly enhanced activity under RSI and HTHH in comparison with the control plants. The silencing of CaZNF830 by virus induced gene silencing (VIGS) significantly compromised pepper immunity against RSI with enhanced growth of Ralstonia solanacearum in pepper plants. Silencing of CaZNF830 also impaired tolerance to HTHH coupled with decreased expression levels of immunity and thermo-tolerance associated marker genes including CaHIR1, CaNPR1, CaPR1, CaABR1 and CaHSP24. By contrast, the transient over-expression of CaZNF830 in pepper leaves by infiltration of GV3101 cells containing 35S::CaZNF830-HA induced HR mimic cell death, H2O2 accumulation and activated the transcriptions of the tested defense-relative or thermo-tolerance associated marker genes. RT-PCR and immune-blotting assay confirmed the stable expression of HA-tagged CaZNF830 mRNA and protein in pepper. All these results suggest that CaZNF830 acts as a positive regulator of plant immunity against RSI or tolerance to HTHH, it is induced by RSI or HTHH and consequently activate pepper immunity against RSI or tolerance to HTHH by directly or indirectly transcriptional modulation of many defense-linked genes.