Identifying Bottle Gourd Genes Responsive to the Infection of Papaya Ringspot Virus

Bottle gourd is an important crop that has helped both ancient and modern civilizations thrive. Besides its applications in medicine, musical instruments, and containers, bottle gourd fruit has relatively high nutritional value in its early stages of development, establishing it as a major food staple in developing countries. Another essential property of bottle gourd lies in its ability to be used as rootstock for grafting to other cucurbit crops. By exploiting this property, farmers can drastically increase their annual yield because grafting increases the scion’s tolerance to abiotic and biotic stresses. That being said, it is necessary that we develop novel methods of increasing pathogen resistance in bottle gourd, as doing so will not only improve the uses that we already have for this crop, but it will also magnify the benefit to other cucurbit crops. On the other hand, Papaya ringspot virus (PRSV) is a major limiting factor of cucurbit production, and it effectively thwarts the benefits of bottle gourd by inhibiting development in young plants and deforming the fruit of mature specimens. To gain insight into the molecular mechanisms underlying bottle gourd resistance to PRSV, we assessed global transcriptome changes in both resistant (USVL5) and susceptible (USVL10) bottle gourd accessions upon infection at 7, 14, and 21 DPI (days post infection). At the beginning of our study, we obtained RNA-Seq transcriptome data, which were subsequently cleaned through the removal of adaptors, low-quality sequences, virus contamination, and rRNA sequences. We then mapped each read to the reference bottle gourd genome, and differentially expressed genes (DEGs) were then identified at each time point for each accession based on the read counts obtained from alignments to the reference genome. Ultimately, we found more DEGs in the susceptible accession upon the PRSV infection than in the resistant accession, especially at 7 DPI. Furthermore, by analyzing the expression patterns of DEGs in the two genotypes and at different days post infection, we identified four clusters of genes with different expression characteristics. Gene Ontology (GO) term enrichment analyses were carried out on these clusters, and significantly overrepresented biological processes and molecular functions were identified. Our results indicated that while genes involved in DNA replication, protein refolding, and stress responses were upregulated in Clusters 1 and 2, genes related to photosynthesis and hormone production were downregulated in Clusters 3 and 4. Interestingly, two genes encoding Argonaute (AGO) proteins, which are essential to antiviral RNA silencing, were upregulated in Clusters 1 and 2, respectively. RNA silencing is one of the known mechanisms underlying plant resistance to virus, and further investigation into the genes involved in the antiviral RNA silencing pathway, such as those encoding RNA-dependent RNA polymerase (RDR), Dicer-like (DCL) proteins, and AGO proteins, showed that their expression levels all started out high in the USVL10 accession at 7 DPI. Overall, our study provides greater insight into the responses of bottle gourd accessions with different levels of resistance to PRSV, which could be applied to future crop improvement through breeding and genetic modification.

My Experience

At the beginning of the PGRP internship, I walked into BTI’s facility with the hope that I could successfully combine my interests in computer science and biology while also learning more about research, data analysis, and lab etiquette. Now, as my internship is drawing to a close, I can state – with a degree of certainty – that my experience at BTI has taken my understanding of all these areas to another level. Although I had conducted many research projects at school, my work at BTI exposed myself to a professional lab setting unlike any that I had worked in before. Challenges that I faced, such as adapting to other programming languages that I had no prior experience with, were admittedly daunting, but overcoming them was truly the biggest step in contributing to my growth as a member of the Fei Lab. All in all, working at BTI has given me a higher level of appreciation for conducting lab research, and in the future, I hope to integrate the skills that I have learned this summer in a manner that not only benefits myself, but also the community as a whole.

As a final note, I would like to thank my mentor, Shan Wu, for guiding me through the research process, as well as Dr. Fei for giving me such an incredible opportunity to work at BTI. In addition, I would also like to thank Mr. Dempsey, Mrs. Pawlowska, and Mrs. McDonald, for they are the individuals who continually motivate me to work hard and follow my dreams.