Structural variants involved in high-altitude adaptation detected using single-molecule long-read sequencing.

Structural variants (SVs), accounting for a larger fraction of the genome than SNPs/InDels, are an important pool of genetic variation, enabling environmental adaptations. Here, we perform long-read sequencing data of 320 Tibetan and Han samples and show that SVs are highly involved in high-altitude adaptation. We expand the landscape of global SVs, apply robust models of selection and population differentiation combining SVs, SNPs and InDels, and use epigenomic analyses to predict enhancers, target genes and biological functions. We reveal diverse Tibetan-specific SVs affecting the regulatory circuitry of biological functions, including the hypoxia response, energy metabolism and pulmonary function. We find a Tibetan-specific deletion disrupts a super-enhancer and downregulates EPAS1 using enhancer reporter, cellular knock-out and DNA pull-down assays. Our study expands the global SV landscape, reveals the role of gene-regulatory circuitry rewiring in human adaptation, and illustrates the diverse functional roles of SVs in human biology.

Oncogenic RAS promotes MYC protein stability by upregulating the expression of the inhibitor of apoptosis protein family member Survivin.

The small GTPase KRAS is frequently mutated in pancreatic cancer and its cooperation with the transcription factor MYC is essential for malignant transformation. The key to oncogenic KRAS and MYC working together is the stabilization of MYC expression due to KRAS activating the extracellular signal-regulated kinase 1/2, which phosphorylates MYC at serine 62 (Ser 62). This prevents the proteasomal degradation of MYC while enhancing its transcriptional activity. Here, we identify how this essential signaling connection between oncogenic KRAS and MYC expression is mediated by the inhibitor of apoptosis protein family member Survivin. This discovery stemmed from our finding that Survivin expression is downregulated upon treatment of pancreatic cancer cells with the KRASG12C inhibitor Sotorasib. We went on to show that oncogenic KRAS increases Survivin expression by activating extracellular signal-regulated kinase 1/2 in pancreatic cancer cells and that treating the cells either with siRNAs targeting Survivin or with YM155, a small molecule that potently blocks Survivin expression, downregulates MYC and strongly inhibited their growth. We further determined that Survivin protects MYC from degradation by blocking autophagy, which then prevents cellular inhibitor of protein phosphatase 2A from undergoing autophagic degradation. Cellular inhibitor of protein phosphatase 2A, by inhibiting protein phosphatase 2A, helps to maintain MYC phosphorylation at Ser 62, thereby ensuring its cooperation with oncogenic KRAS in driving cancer progression. Overall, these findings highlight a novel role for Survivin in mediating the cooperative actions of KRAS and MYC during malignant transformation and raise the possibility that targeting Survivin may offer therapeutic benefits against KRAS-driven cancers.

The GGC repeat expansion in NOTCH2NLC is associated with oculopharyngodistal myopathy type 3.

Oculopharyngodistal myopathy (OPDM) is an adult-onset neuromuscular disease characterized by progressive ocular, facial, pharyngeal and distal limb muscle involvement. Trinucleotide repeat expansions in LRP12 or GIPC1 were recently reported to be associated with OPDM. However, a significant portion of OPDM patients have unknown genetic causes. In this study, long-read whole-genome sequencing and repeat-primed PCR were performed and we identified GGC repeat expansions in the NOTCH2NLC gene in 16.7% (4/24) of a cohort of Chinese OPDM patients, designated as OPDM type 3 (OPDM3). Methylation analysis indicated that methylation levels of the NOTCH2NLC gene were unaltered in OPDM3 patients, but increased significantly in asymptomatic carriers. Quantitative real-time PCR analysis indicated that NOTCH2NLC mRNA levels were increased in muscle but not in blood of OPDM3 patients. Immunofluorescence on OPDM muscle samples and expressing mutant NOTCH2NLC with (GGC)69 repeat expansions in HEK293 cells indicated that mutant NOTCH2NLC-polyglycine protein might be a major component of intranuclear inclusions, and contribute to toxicity in cultured cells. In addition, two RNA-binding proteins, hnRNP A/B and MBNL1, were both co-localized with p62 in intranuclear inclusions in OPDM muscle samples. These results indicated that a toxic protein gain-of-function mechanism and RNA gain-of-function mechanism may both play a vital role in the pathogenic processes of OPDM3. This study extended the spectrum of NOTCH2NLC repeat expansion-related diseases to a predominant myopathy phenotype presenting as OPDM, and provided evidence for possible pathogenesis of these diseases.

Preferential gene retention increases the robustness of cold regulation in Brassicaceae and other plants after polyploidization.

Cold stress profoundly affects plant growth and development and is a key factor affecting the geographic distribution and evolution of plants. Plants have evolved adaptive mechanisms to cope with cold stress. Here, through the genomic analysis of Arabidopsis, three Brassica species and 17 other representative species, we found that both cold-related genes (CRGs) and their collinearity were preferentially retained after polyploidization followed by genome instability, while genome-wide gene sets exhibited a variety of other expansion mechanisms. The cold-related regulatory network was increased in Brassicaceae genomes, which were recursively affected by polyploidization. By combining our findings regarding the selective retention of CRGs from this ecological genomics study with the available knowledge of cold-induced chromosome doubling, we hypothesize that cold stress may have contributed to the success of polyploid plants through both increasing polyploidization and selectively maintaining CRGs during evolution. This hypothesis requires further biological and ecological exploration to obtain solid supporting evidence, which will potentially contribute to understanding the generation of polyploids and to the field of ecological genomics.

An Overlooked Paleotetraploidization in Cucurbitaceae.

Cucurbitaceae plants are of considerable biological and economic importance, and genomes of cucumber, watermelon, and melon have been sequenced. However, a comparative genomics exploration of their genome structures and evolution has not been available. Here, we aimed at performing a hierarchical inference of genomic homology resulted from recursive paleopolyploidizations. Unexpectedly, we found that, shortly after a core-eudicot-common hexaploidy, a cucurbit-common tetraploidization (CCT) occurred, overlooked by previous reports. Moreover, we characterized gene loss (and retention) after these respective events, which were significantly unbalanced between inferred subgenomes, and between plants after their split. The inference of a dominant subgenome and a sensitive one suggested an allotetraploid nature of the CCT. Besides, we found divergent evolutionary rates among cucurbits, and after doing rate correction, we dated the CCT to be 90-102 Ma, likely common to all Cucurbitaceae plants, showing its important role in the establishment of the plant family.

Alignment of Common Wheat and Other Grass Genomes Establishes a Comparative Genomics Research Platform.

Grass genomes are complicated structures as they share a common tetraploidization, and particular genomes have been further affected by extra polyploidizations. These events and the following genomic re-patternings have resulted in a complex, interweaving gene homology both within a genome, and between genomes. Accurately deciphering the structure of these complicated plant genomes would help us better understand their compositional and functional evolution at multiple scales. Here, we build on our previous research by performing a hierarchical alignment of the common wheat genome vis-à-vis eight other sequenced grass genomes with most up-to-date assemblies, and annotations. With this data, we constructed a list of the homologous genes, and then, in a layer-by-layer process, separated their orthology, and paralogy that were established by speciations and recursive polyploidizations, respectively. Compared with the other grasses, the far fewer collinear outparalogous genes within each of three subgenomes of common wheat suggest that homoeologous recombination, and genomic fractionation should have occurred after its formation. In sum, this work contributes to the establishment of an important and timely comparative genomics platform for researchers in the grass community and possibly beyond. Homologous gene list can be found in Supplemental material.

Two Highly Similar Poplar Paleo-subgenomes Suggest an Autotetraploid Ancestor of Salicaceae Plants.

As a model plant to study perennial trees in the Salicaceae family, the poplar (Populus trichocarpa) genome was sequenced, revealing recurrent paleo-polyploidizations during its evolution. A comparative and hierarchical alignment of its genome to a well-selected reference genome would help us better understand poplar's genome structure and gene family evolution. Here, by adopting the relatively simpler grape (Vitis vinifera) genome as reference, and by inferring both intra- and inter-genomic gene collinearity, we produced a united alignment of these two genomes and hierarchically distinguished the layers of paralogous and orthologous genes, as related to recursive polyploidizations and speciation. We uncovered homologous blocks in the grape and poplar genomes and also between them. Moreover, we characterized the genes missing and found that poplar had two considerably similar subgenomes (≤0.05 difference in gene deletion) produced by the Salicaceae-common tetraploidization, suggesting its autotetraploid nature. Taken together, this work provides a timely and valuable dataset of orthologous and paralogous genes for further study of the genome structure and functional evolution of poplar and other Salicaceae plants.

Hierarchically Aligning 10 Legume Genomes Establishes a Family-Level Genomics Platform.

Mainly due to their economic importance, genomes of 10 legumes, including soybean (Glycine max), wild peanut (Arachis duranensis and Arachis ipaensis), and barrel medic (Medicago truncatula), have been sequenced. However, a family-level comparative genomics analysis has been unavailable. With grape (Vitis vinifera) and selected legume genomes as outgroups, we managed to perform a hierarchical and event-related alignment of these genomes and deconvoluted layers of homologous regions produced by ancestral polyploidizations or speciations. Consequently, we illustrated genomic fractionation characterized by widespread gene losses after the polyploidizations. Notably, high similarity in gene retention between recently duplicated chromosomes in soybean supported the likely autopolyploidy nature of its tetraploid ancestor. Moreover, although most gene losses were nearly random, largely but not fully described by geometric distribution, we showed that polyploidization contributed divergently to the copy number variation of important gene families. Besides, we showed significantly divergent evolutionary levels among legumes and, by performing synonymous nucleotide substitutions at synonymous sites correction, redated major evolutionary events during their expansion. This effort laid a solid foundation for further genomics exploration in the legume research community and beyond. We describe only a tiny fraction of legume comparative genomics analysis that we performed; more information was stored in the newly constructed Legume Comparative Genomics Research Platform (www.legumegrp.org).

Comparative Genomics Analysis of Rice and Pineapple Contributes to Understand the Chromosome Number Reduction and Genomic Changes in Grasses.

Rice is one of the most researched model plant, and has a genome structure most resembling that of the grass common ancestor after a grass common tetraploidization ∼100 million years ago. There has been a standing controversy whether there had been five or seven basic chromosomes, before the tetraploidization, which were tackled but could not be well solved for the lacking of a sequenced and assembled outgroup plant to have a conservative genome structure. Recently, the availability of pineapple genome, which has not been subjected to the grass-common tetraploidization, provides a precious opportunity to solve the above controversy and to research into genome changes of rice and other grasses. Here, we performed a comparative genomics analysis of pineapple and rice, and found solid evidence that grass-common ancestor had 2n = 2x = 14 basic chromosomes before the tetraploidization and duplicated to 2n = 4x = 28 after the event. Moreover, we proposed that enormous gene missing from duplicated regions in rice should be explained by an allotetraploid produced by prominently divergent parental lines, rather than gene losses after their divergence. This means that genome fractionation might have occurred before the formation of the allotetraploid grass ancestor.