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The most important role of variant pathogenicity predictors is to identify the disease-phenotype causative variant in studying monogenic diseases. In the last decade, machine-learning based predictors exhibited a relatively accurate performance for distinguishing the pathogenic variants and contributed a significant role for all disease-spectrums. Yet, few predictors can investigate the phenotypic significance of variants. Here we presented a phenotype-specific framework aimed to directly point out the phenotypic significance of predicted candidates, and showed its advancing performance in eye abnormalities. By training on eye-abnormalities causative variants, our method presented 96.2% accuracy, 96.1% precision, 93.4% recall for pathogenicity identification. Inconsistent with the modeling performance, identifying the single phenotype-causative variant from various sequencing variants is challenging for all predictors. Underlying the phenotype-oriented, our method significantly promoted the precision and reduced the cost for identifying the single causative variant from thousands of candidates. These advances highlight the significance of the phenotype-specific training method for studying disease.

Hereditary spastic paraplegia (HSP) is a neurodegenerative disease characterized by progressive weakness and spasticity of the lower limbs, in complicated forms, with additional neurological signs. To identify the genotype and characterize the phenotype in a Chinese HSP family, ten subjects from the family were examined through detailed clinical evaluations, auxiliary examinations and genetic tests. Using a combined approach of whole-exome sequencing and candidate mutation validation, we identified novel compound heterozygous mutations in the SPG11 gene of the patients as follows: a nonsense mutation c.6856C>T (p.R2286X) in exon 38 and a deletion mutation c.2863delG (p.Glu955Lysfs*8) in exon 16. Both mutations co-segregated with the phenotype in this family and were absent in 100 normal Chinese individuals. Our finding suggests that the novel compound heterozygous mutations in SPG11 are associated with HSP. We were able to assess the future risk of HSP in healthy younger family members using genetic detection, and provide prenatal diagnoses for the family members. Furthermore, to some extent, this new finding enriches the information on SPG11 and may provide a new basis for the genetic diagnosis of HSP.

Olmsted syndrome (OS) is a rare congenital disorder characterized by palmoplantar and periorificial keratoderma, alopecia in most cases, and severe itching. The genetic basis for OS remained unidentified. Using whole-exome sequencing of case-parents trios, we have identified a de novo missense mutation in TRPV3 that produces p.Gly573Ser in an individual with OS. Nucleotide sequencing of five additional affected individuals also revealed missense mutations in TRPV3 (which produced p.Gly573Ser in three cases and p.Gly573Cys and p.Trp692Gly in one case each). Encoding a transient receptor potential vanilloid-3 cation channel, TRPV3 is primarily expressed in the skin, hair follicles, brain, and spinal cord. In transfected HEK293 cells expressing TRPV3 mutants, much larger inward currents were recorded, probably because of the constitutive opening of the mutants. These gain-of-function mutations might lead to elevated apoptosis of keratinocytes and consequent skin hyperkeratosis in the affected individuals. Our findings suggest that TRPV3 plays essential roles in skin keratinization, hair growth, and possibly itching sensation in humans and selectively targeting TRPV3 could provide therapeutic potential for keratinization or itching-related skin disorders.

Promoters that have been widely used for both basic research and biotechnological application in plants are generally unidirectional. Here we describe a strategy to bi-directionalize the vascular-specific expression grp1.8 promoter (here named GRPp) and 4CL1 promoter (here named 4CL1p) so that one promoter can direct the vascular-specific expression of two genes, one on each end of the promoter. The minimal promoter (35Smini or GRP mini), when fused at the 5' end of the specific expression promoter (GRPp or 4CL1p) to form bi-directional promoter (35Smini-GRPp, 35Smini-4CL1p or GRPmini-GRPp), was able to direct expression of the glucuronidase (gus) and green fluorescent protein (gfp) gene in all independent transgenic tobacco lines. Stable expression of gusA and gfp genes in transgenic plants was analyzed by histochemical staining for GUS and fluorescence microscopic observation under UV for GFP in transgenic plants. The remarkable transcript levels of GFP and GUS were detected by real-time PCR in independent transgenic tobacco lines. Their vascular-specific bi-directional promoters should be used to vascular-specific expression several functional genes in transgenic plants simultaneously.