Deep learning-based characterization and redesign of major potato tuber storage protein.

Potato is one of the most important crops worldwide, to feed a fast-growing population. In addition to providing energy, fiber, vitamins, and minerals, potato storage proteins are considered as one of the most valuable sources of non-animal proteins due to their high essential amino acid (EAA) index. However, low tuber protein content and limited knowledge about potato storage proteins restrict their widespread utilization in the food industry. Here, we report a proof-of-concept study, using deep learning-based protein design tools, to characterize the biological and chemical characteristics of patatins, the major potato storage proteins. This knowledge was then employed to design multiple cysteines on the patatin surface to build polymers linked by disulfide bonds, which significantly improved viscidity and nutrient of potato flour dough. Our study shows that deep learning-based protein design strategies are efficient to characterize and to create novel proteins for future food sources.

Genome-wide identification and gene expression analysis of the 14-3-3 gene family in potato (Solanum tuberosum L.).

BACKGROUND: 14-3-3 proteins are essential in regulating various biological processes and abiotic stress responses in plants. Although 14-3-3 proteins have been studied in model plants such as Arabidopsis thaliana and Oryza sativa, there is a lack of research on the 14-3-3 gene family in potatoes (Solanum tuberosum L.). RESULTS: A total of 18 14-3-3 genes encoding proteins containing a typical conserved PF00244 domain were identified by genome-wide analysis in potatoes. The St14-3-3 gene family members were unevenly distributed across the chromosomes, and gene structure analysis showed that gene length and intron number varied greatly among the members. Phylogenetic analysis of 14-3-3 proteins in potatoes and other plant species showed that they could be divided into two distinct groups (ε and non-ε). Members in the ε group tended to have similar exon-intron structures and conserved motif patterns. Promoter sequence analysis showed that the St14-3-3 gene promoters contained multiple hormone-, stress-, and light-responsive cis-regulatory elements. Synteny analysis suggested that segmental duplication events contributed to the expansion of the St14-3-3 gene family in potatoes. The observed syntenic relationships between some 14-3-3 genes from potato, Arabidopsis, and tomato suggest that they evolved from a common ancestor. RNA-seq data showed that St14-3-3 genes were expressed in all tissues of potatoes but that their expression patterns were different. qRT-PCR assays revealed that the expression levels of nearly all tested St14-3-3 genes were affected by drought, salt, and low-temperature stresses and that different St14-3-3 genes had different responses to these stresses. CONCLUSIONS: In summary, genome-wide identification, evolutionary, and expression analyses of the 14-3-3 gene family in potato were conducted. These results provide important information for further studies on the function and regulation of St14-3-3 gene family members in potatoes.

Fine Mapping and Candidate Gene Prediction of Tuber Shape Controlling Ro Locus Based on Integrating Genetic and Transcriptomic Analyses in Potato.

Tuber shape is one of the most important quality traits in potato appearance. Since poor or irregular shape results in higher costs for processing and influences the consumers' willingness to purchase, breeding for shape uniformity and shallow eye depth is highly important. Previous studies showed that the major round tuber shape controlling locus, the Ro locus, is located on chromosome 10. However, fine mapping and cloning of tuber shape genes have not been reported. In this study, the analyses of tissue sectioning and transcriptome sequencing showed that the developmental differences between round and elongated tuber shapes begin as early as the hook stage of the stolon. To fine map tuber shape genes, a high-density genetic linkage map of the Ro region on chromosome 10 based on a diploid segregating population was constructed. The total length of the genetic linkage map was 25.8 cM and the average marker interval was 1.98 cM. Combined with phenotypic data collected from 2014 to 2017, one major quantitative trait locus (QTL) for tuber shape was identified, which explained 61.7-72.9% of the tuber shape variation. Through the results of genotyping and phenotypic investigation of recombinant individuals, Ro was fine mapped in a 193.43 kb interval, which contained 18 genes. Five candidate genes were preliminarily predicted based on tissue sections and transcriptome sequencing. This study provides an important basis for cloning Ro gene(s).

Genome-wide simple sequence repeat markers in potato: abundance, distribution, composition, and polymorphism.

Simple sequence repeats (SSRs) are important sources of genetic diversity and are widely used as markers in genetics and molecular breeding. In this study, we examined four potato genomes of DM1-3 516 R44 (DM) from Solanum phureja, RH89039-16 (RH) from Solanum tuberosum, M6 from Solanum chacoense and Solanum commersonii to determine SSR abundance and distribution and develop a larger list of polymorphic markers for a potentially wide range of uses for the potato community. A total of 1,734,619 SSRs were identified across the four genomes with an average of 433,655 SSRs per genome and 2.31kb per SSR. The most abundant repeat units for mono-, di-, tri-, and tetra-nucleotide SSRs were (A/T)n, (AT/AT)n, (AAT/ATT)n, and (ATAT/ATAT)n, respectively. The SSRs were most abundant (78.79%) in intergenic regions and least abundant (3.68%) in untranslated regions. On average, 168,069 SSRs with unique flanking sequences were identified in the four genomes. Further, we identified 16,245 polymorphic SSR markers among the four genomes. Experimental validation confirmed 99.69% of tested markers could generate target bands. The high-density potato SSR markers developed in this study will undoubtedly facilitate the application of SSR markers for genetic research and marker-pyramiding in potato breeding.

New Findings on the Resistance Mechanism of an Elite Diploid Wild Potato Species JAM1-4 in Response to a Super Race Strain of Phytophthora infestans.

Late blight is a devastating potato disease worldwide, caused by Phytophthora infestans. The P. infestans strain 2013-18-306 from Yunnan is a "supervirulent race" that overcomes all 11 known late blight resistance genes (R1 to R11) from Solanum demissum. In a previous study, we identified a diploid wild-type potato JAM1-4 (S. jamesii) with high resistance to 2013-18-306. dRenSeq analysis indicated the presence of novel R genes in JAM1-4. RNA-Seq was used to analyze the late blight resistance response genes and defense regulatory mechanisms of JAM1-4 against 2013-18-306. Gene ontology enrichment and KEGG pathway analysis showed that many disease-resistant pathways were significantly enriched. Analysis of differentially expressed genes (DEGs) revealed an active disease resistance mechanism of JAM1-4, and the essential role of multiple signal transduction pathways and secondary metabolic pathways comprised of SA-JA-ET in plant immunity. We also found that photosynthesis in JAM1-4 was inhibited to promote the immune response. Our study reveals the pattern of resistance-related gene expression in response to a super race strain of potato late blight and provides a theoretical basis for further exploration of potato disease resistance mechanisms, discovery of new late blight resistance genes, and disease resistance breeding.

Comparative Transcriptome Profiling Reveals Compatible and Incompatible Patterns of Potato Toward Phytophthora infestans.

Late blight, caused by Phytophthora infestans (P. infestans), is a devastating disease in potato worldwide. Our previous study revealed that the Solanum andigena genotype 03112-233 is resistant to P. infestans isolate 90128, but susceptible to the super race isolate, CN152. In this study, we confirmed by diagnostic resistance gene enrichment sequencing (dRenSeq) that the resistance of 03112-233 toward 90128 is most likely based on a distinct new R gene(s). To gain an insight into the mechanism that governs resistance or susceptibility in 03112-223, comparative transcriptomic profiling analysis based on RNAseq was initiated. Changes in transcription at two time points (24 h and 72 h) after inoculation with isolates 90128 or CN152 were analyzed. A total of 8,881 and 7,209 genes were differentially expressed in response to 90128 and CN152, respectively, and 1,083 differentially expressed genes (DEGs) were common to both time points and isolates. A substantial number of genes were differentially expressed in an isolate-specific manner with 3,837 genes showing induction or suppression following infection with 90128 and 2,165 genes induced or suppressed after colonization by CN152. Hierarchical clustering analysis suggested that isolates with different virulence profiles can induce different defense responses at different time points. Further analysis revealed that the compatible interaction caused higher induction of susceptibility genes such as SWEET compared with the incompatible interaction. The salicylic acid, jasmonic acid, and abscisic acid mediated signaling pathways were involved in the response against both isolates, while ethylene and brassinosteroids mediated defense pathways were suppressed. Our results provide a valuable resource for understanding the interactions between P. infestans and potato.

Discovery of novel oxoindolin derivatives as atypical dual inhibitors for DNA Gyrase and FabH.

The antibacterial agents and therapies today are facing serious problems such as drug resistance. Introducing dual inhibiting effect is a valid approach to solve this trouble and bring advantages including wide adaptability, favorable safety and superiority of combination. We started from potential DNA Gyrase inhibitory backbone isatin to develop oxoindolin derivatives as atypical dual Gyrase (major) and FabH (assistant) inhibitors via a two-round screening. Aiming at blocking both duplication (Gyrase) and survival (FabH), most of synthesized compounds indicated potency against Gyrase and some of them inferred favorable inhibitory effect on FabH. The top hit I18 suggested comparable Gyrase inhibitory activity (IC50 = 0.025 µM) and antibacterial effect with the positive control Novobiocin (IC50 = 0.040 µM). FabH inhibitory activity (IC50 = 5.20 µM) was also successfully introduced. Docking simulation hinted possible important interacted residues and binding patterns for both target proteins. Adequate Structure-Activity Relation discussions provide the future orientations of modification. With high potency, low initial toxicity and dual inhibiting strategy, advanced compounds with therapeutic methods will be developed for clinical application.

Discovery and development of novel rhodanine derivatives targeting enoyl-acyl carrier protein reductase.

A series of rhodanine derivatives RB1-RB23 were synthesized through a two-round screening. Their Mycobacterial tuberculosis (Mtb) InhA inhibitory activity and Mtb growth blocking capability were evaluated. The most potent hit compound RB23 indicated comparable InhA inhibiton (IC50 = 2.55 µM) with the positive control Triclosan (IC50 = 6.14 µM) and Isoniazid (IC50 = 8.29 µM). Its improved growth-blocking effect on Mtb and low toxicity were attractive for further development. The docking simulation revealed the possible binding pattern of this series and picked the key interacted residues as Ser20, Phe149, Lys165 and Thr196. The 3D-QSAR model visualized the SAR discussion and hinted new information. Modifying the surroundings near rhodanine moiety might be promising attempts in later investigations.

Transcriptome Profiling Reveals Effects of Drought Stress on Gene Expression in Diploid Potato Genotype P3-198.

Potato (Solanum tuberosum L.) is one of the three most important food crops worldwide; however, it is strongly affected by drought stress. The precise molecular mechanisms of drought stress response in potato are not very well understood. The diploid potato genotype P3-198 has been verified to be highly resistant to drought stress. Here, a time-course experiment was performed to identify drought resistance response genes in P3-198 under polyethylene glycol (PEG)-induced stress using RNA-sequencing. A total of 1665 differentially expressed genes (DEGs) were specifically identified, and based on gene ontology and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis, the transcription factor activity, protein kinase activity, and the plant hormone signal transduction process were significantly enriched. Annotation revealed that these DEGs mainly encode transcription factors, protein kinases, and proteins related to redox regulation, carbohydrate metabolism, and osmotic adjustment. In particular, genes encoding abscisic acid (ABA)-dependent signaling molecules were significantly differentially expressed, which revealed the important roles of the ABA-dependent signaling pathway in the early response of P3-198 to drought stress. Quantitative real-time PCR experimental verification confirmed the differential expression of genes in the drought resistance signaling pathway. Our results provide valuable information for understanding potato drought-resistance mechanisms, and also enrich the gene resources available for drought-resistant potato breeding.

Tracking disease resistance deployment in potato breeding by enrichment sequencing.

Following the molecular characterisation of functional disease resistance genes in recent years, methods to track and verify the integrity of multiple genes in varieties are needed for crop improvement through resistance stacking. Diagnostic resistance gene enrichment sequencing (dRenSeq) enables the high-confidence identification and complete sequence validation of known functional resistance genes in crops. As demonstrated for tetraploid potato varieties, the methodology is more robust and cost-effective in monitoring resistances than whole-genome sequencing and can be used to appraise (trans) gene integrity efficiently. All currently known NB-LRRs effective against viruses, nematodes and the late blight pathogen Phytophthora infestans can be tracked with dRenSeq in potato and hitherto unknown polymorphisms have been identified. The methodology provides a means to improve the speed and efficiency of future disease resistance breeding in crops by directing parental and progeny selection towards effective combinations of resistance genes.

Mapping and QTL Analysis of Early-Maturity Traits in Tetraploid Potato (Solanum tuberosum L.).

Early maturity is one of the most important agronomical traits in potato breeding. To identify the DNA segment that codes for early maturity, a tetraploid potato segregation population of "Zhongshu 19" × "Zhongshu 3" was genetically analyzed, using a combination of high throughput simplified genome sequencing (2b-RAD) and bulked segregant analysis (BSA). The DNA segment related to the early-maturity trait was identified at the 3.7~4.2 Mb locus on the short arm of chromosome 5. Eight molecular markers were developed, of which five were closely linked to the early-maturity trait loci. Additionally, 42 simple sequence repeats (SSR) markers were constructed based on the reference sequence of Solanum tuberosum group Phureja DM1-3 516 R44 (DM). Using the TetraploidMap software, the linkage map of chromosome 5 was constructed with 50 markers. The total map length was 172 centiMorgan (cM), with an average genetic distance of 3.44 cM. Correlating molecular and phenotypic data of the segregating population, the mapped Quantitative Trait Loci (QTL) on the short arm of chromosome 5 contributed to 33.55% of the early-maturity phenotype. The early-maturity QTL was located at 84 cM, flanked by the SSR5-85-1 and SCAR5-8 markers. The QTL was fine-mapped to 471 kb. Using DNA sequence annotation, 34 genes were identified in this region, 12 of them with unknown function. Among the other 22 annotated genes, E3 ubiquitin ligase gene PUB14 could be related to maturity and regulation of tuber formation. The constructed QTL map is a useful basic tool for the cloning of early-maturity related genes in tetraploid potatoes.

Genome sequence and analysis of the tuber crop potato.

Potato (Solanum tuberosum L.) is the world's most important non-grain food crop and is central to global food security. It is clonally propagated, highly heterozygous, autotetraploid, and suffers acute inbreeding depression. Here we use a homozygous doubled-monoploid potato clone to sequence and assemble 86% of the 844-megabase genome. We predict 39,031 protein-coding genes and present evidence for at least two genome duplication events indicative of a palaeopolyploid origin. As the first genome sequence of an asterid, the potato genome reveals 2,642 genes specific to this large angiosperm clade. We also sequenced a heterozygous diploid clone and show that gene presence/absence variants and other potentially deleterious mutations occur frequently and are a likely cause of inbreeding depression. Gene family expansion, tissue-specific expression and recruitment of genes to new pathways contributed to the evolution of tuber development. The potato genome sequence provides a platform for genetic improvement of this vital crop.