Chlorophyll fluorescence imaging for monitoring effects of Heterobasidion parviporum small secreted protein induced cell death and in planta defense gene expression

Abstract

Heterobasidion parviporum Niemelä & Korhonen is a necrotrophic fungal pathogen of Norway spruce (Picea abies). H. parviporum genome encodes numerous necrotrophic small secreted proteins (SSP) which might be important for promoting and sustaining the disease development. However, their transcriptional dynamics and plant defense response during infection are largely unknown. In this study, we identified a necrotrophic SSP named HpSSP35.8 and its coding gene was highly expressed in the pre-symptomatic phase of host infection. We explored the impact of HpSSP35.8 on Nicotiana benthamiana using Agrobacterium-mediated transient expression system under visible spectrum RGB imaging and chlorophyll fluorescence imaging. The results showed that HpSSP35.8 triggered a form of SSP-associated programmed cell death, accompanied by a decrease in the plant photosynthetic activity. Defense-related genes including WRKY12, ethylene response factor (ERF1α) and a chitinase gene PR4 were up-regulated in both HpSSP35.8-N. benthamiana interaction and H. parviporum-Norway spruce pathosystem. It highlighted the potential for the use of chlorophyll fluorescence imaging approach for monitoring indirect effects of SSP as well as for selection of other potential effector-like protein candidates.

The Conifer Root and Stem Rot Pathogen (Heterobasidion parviporum): Effectome Analysis and Roles in Interspecific Fungal Interactions

“…HaSSP16, homolog of HpSSP3.534 in H. annosum, showed low transcript level in necrotrophic conditions, but high transcript level in saprotrophic growth in wood [32]. HpSSP35.8 probably had a role in the disease process based on its high expression in the presymptomatic phase of seedling infection, and it induced SSP-associated programmed cell death in Nicotiana benthamiana [33]. The expression level of HpSSP33.48 was higher than that of HpSSP33.91, especially in SPORE, which revealed that they are probably different in function.…”

The dark septate endophyte Phialocephala sphaeroides suppresses conifer pathogen transcripts and promotes root growth of Norway spruce

“…2020 ) in both groups of samples. The expression of effector HpSSP35.8 gene, which induces cell death and tree defence responses ( Wen, Raffaello, et al. 2019 ), was kept at comparable levels in both groups of samples.…”

Effectors of Plant Necrotrophic Fungi

“…Recent bioinformatic studies have revealed repertoires of small secreted proteins (SSPs) in several phytopathogens that might contribute to virulence, and many of them are cysteine-rich, small-sized proteins without known protein domains (Wen et al, 2019;Denton-Giles et al, 2020). Predicted effectors with no known domains from a transcriptome study of S. sclerotiorum were recently screened for cell death-inducing activity (Seifbarghi et al, 2020).…”

“…In the conifer pathogen Heterobasidion parviporum, a SSP without any known protein domains, HpSSP35.8, was identified as a necrotrophic effector protein secreted during infection (Wen et al, 2019). Overexpression of HpSSP35.8 induces a strong cell death phenotype and activates PR genes, the "WRKY" transcription factors, genes involved in JA/ET-signaling, and chitinase genes, all of which are known defense responses (Wen et al, 2019). This response suggests that the cell death observed is an HR-like response, though HpSSP35.8 also had significant effects on chlorophyll and photosynthesis, which could indicate that the cell death observed is truly necrosis.…”