Hyperosmolality and various other stimuli can trigger an increase in cytoplasmic-free calcium concentration ([Ca²⁺]_cyt). Members of the Arabidopsis thaliana (L.) reduced hyperosmolality-gated calcium-permeable channels (OSCA) gene family are reported to be involved in sensing extracellular changes to trigger hyperosmolality-induced [Ca²⁺]_cyt increases and controlling stomatal closure during immune signaling. Wheat (Triticum aestivum L.) is a very important food crop, but there are few studies of its OSCA gene family members. In this study, 42 OSCA members were identified in the wheat genome, and phylogenetic analysis can divide them into four clades. The members of each clade have similar gene structures, conserved motifs, and domains. TaOSCA genes were predicted to be regulated by cis-acting elements such as STRE, MBS, DRE1, ABRE, etc. Quantitative PCR results showed that they have different expression patterns in different tissues. The expression profiles of 15 selected TaOSCAs were examined after PEG (polyethylene glycol), NaCl, and ABA (abscisic acid) treatment. All 15 TaOSCA members responded to PEG treatment, while TaOSCA12/-39 responded simultaneously to PEG and ABA. This study informs research into the biological function and evolution of TaOSCA and lays the foundation for the breeding and genetic improvement of wheat.

Brassinazole-resistant (BZR) family genes encode plant-specific transcription factors (TFs), play essential roles in the regulation of plant growth and development, and have multiple stress-resistance functions. Nicotiana benthamiana is a model plant widely used in basic research. However, members of the BZR family in N. benthamiana have not been identified, and little is known about their function in abiotic stress. In this study, a total of 14 BZR members were identified in the N. benthamiana genome, which could be divided into four groups according to a phylogenetic tree. NbBZRs have similar exon-intron structures and conserved motifs, and may be regulated by cis-acting elements such as STRE, TCA, and ARE, etc. Organ-specific expression analysis showed that NbBZR members have different and diverse expression patterns in different tissues, and most of the members are expressed in roots, stems, and leaves. The analysis of the expression patterns in response to different abiotic stresses showed that all the tested NbBZR members showed a significant down-regulation after drought treatment. Many NbBZR genes also responded in various ways to cold, heat and salt stress treatments. The results imply that NbBZRs have multiple functions related to stress resistance.

Quantitative real-time PCR (RT-qPCR) is a widely used method for studying alterations in gene expression upon infections caused by diverse pathogens such as viruses. Positive-sense single-stranded (ss(+)) RNA viruses form a major part of all known plant viruses, and some of them are damaging pathogens of agriculturally important crops. Analysis of gene expression following infection by ss(+) RNA viruses is crucial for the identification of potential anti-viral factors. However, viral infections are known to globally affect gene expression and therefore selection and validation of reference genes for RT-qPCR is particularly important. In this study, the expression of commonly used reference genes for RT-qPCR was studied in Nicotiana benthamiana following single infection by 11 ss(+) RNA viruses, including five tobamoviruses, four potyviruses, one potexvirus and one polerovirus. Stability of gene expression was analyzed in parallel by four commonly used algorithms: geNorm, NormFinder, BestKeeper, and Delta CT, and RefFinder was finally used to summarize all the data. The most stably expressed reference genes differed significantly among the viruses, even when those viruses were from the same genus. Our study highlights the importance of the selection and validation of reference genes upon different viral infections.

Tomato brown rugose fruit virus (ToBRFV) is a newly-emerging tobamovirus which was first reported on tomatoes in Israel and Jordan, and which has now spread rapidly in Asia, Europe, North America, and Africa. ToBRFV can overcome the resistance to other tobamoviruses conferred by tomato Tm-1, Tm-2, and Tm-2² genes, and it has seriously affected global crop production. The rapid and comprehensive transcription reprogramming of host plant cells is the key to resisting virus attack, but there have been no studies of the transcriptome changes induced by ToBRFV in tomatoes. Here, we made a comparative transcriptome analysis between tomato leaves infected with ToBRFV for 21 days and those mock-inoculated as controls. A total of 522 differentially expressed genes were identified after ToBRFV infection, of which 270 were up-regulated and 252 were down-regulated. Functional analysis showed that DEGs were involved in biological processes such as response to wounding, response to stress, protein folding, and defense response. Ten DEGs were selected and verified by qRT-PCR, confirming the reliability of the high-throughput sequencing data. These results provide candidate genes or signal pathways for the response of tomato leaves to ToBRFV infection.

Tomato brown rugose fruit virus (ToBRFV) is an emerging, rapidly spreading virus belonging to the genus Tobamovirus which seriously decreases tomato yields. RNA silencing is an evolutionarily conserved antiviral mechanism. In virus-infected plants, virus-derived small interfering RNAs (vsiRNAs) are one of the key components involved in the RNA silencing-based antiviral activity in plants. The main function of vsiRNAs is to target and degrade viral RNA. Studies have found that some vsiRNAs can also target host transcripts, further regulating host responses and symptoms and promoting viral survival and spread. In this study, the vsiRNA profiles of ToBRFV-infected tomato plants were obtained by deep sequencing. VsiRNAs were mainly 21 and 22 nucleotides in length and had a U-bias at the 5′ end. The single-nucleotide resolution profile shows that vsiRNAs exhibit a heterogeneous continuous distribution in the ToBRFV genomic RNA, with hotspot regions on the antisense strand located at the 5′ end of the RdRP and in the coding regions of MP and CP. The presence of vsiRNAs was confirmed in tomato plants infected with ToBRFV through RT-PCR, and GO and KEGG enrichment analyses were performed on the predicted vsiRNA target genes (with an expectation value less than or equal to 2.5). Seven potential target genes were selected for qRT-PCR analysis, confirming that their transcript accumulation significantly decreased in the leaves of tomato plants infected with ToBRFV. These genes may play an important role in the process of viral infection in tomatoes. Our results suggest a role for vsiRNAs in the ToBRFV–tomato interaction.

Chili pepper (Capsicum annuum L.) is a very important vegetable crop, commonly used as a spice or seasoning in various dishes. With the growth of the global population, the demand for chili peppers has also increased exponentially. Tomato brown rugose fruit virus (ToBRFV) is an emerging tobamovirus that has spread to dozens of countries worldwide. Its infection in chili peppers can severely impact yield and quality, posing a significant threat to the chili pepper industry. However, the transcriptional response of chili peppers to ToBRFV infection has not been studied yet. This research utilized RNA-Seq technology to analyze the transcriptional profiles of chili pepper leaves (‘Haonong 11’) 13 days post-infection with ToBRFV or mock treatment, identifying a total of 1468 differentially expressed genes (DEGs), of which 1366 were upregulated and 102 were downregulated. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses indicated that the DEGs were involved in biological processes such as defense response, response to reactive oxygen species, protein folding, and plant-pathogen interaction. Twelve DEGs were validated by RT-qPCR, with their expression trends consistent with the transcriptome data, indicating the reliability of the high-throughput data. Our systematic analysis provides a molecular basis for the response of chili pepper leaves to ToBRFV infection at the transcriptomic level and offers potential candidate genes for further research into the interaction mechanisms between ToBRFV and plant hosts.