An L1 retrotransposon insertion–induced deafness mouse model for studying the development and function of the cochlear stria vascularis

Dysregulation of ion and potential homeostasis in the scala media is the most prevalent cause of hearing loss in mammals. However, it is not well understood how the development and function of the stria vascularis regulates this fluid homeostasis in the scala media. From a mouse genetic screen, we characterize a mouse line, named 299, that displays profound hearing impairment. Histology suggests that 299 mutant mice carry a severe, congenital structural defect of the stria vascularis. The in vivo recording of 299 mice using double-barreled electrodes shows that endocochlear potential is abolished and potassium concentration is reduced to ∼20 mM in the scala media, a stark contrast to the +80 mV endocochlear potential and the 150 mM potassium concentration present in healthy control mice. Genomic analysis revealed a roughly 7-kb-long, interspersed nuclear element (LINE-1 or L1) retrotransposon insertion on chromosome 11. Strikingly, the deletion of this L1 retrotransposon insertion from chromosome 11 restored the hearing of 299 mutant mice. In summary, we characterize a mouse model that enables the study of stria vascularis development and fluid homeostasis in the scala media.

Gibberellins Inhibit Flavonoid Biosynthesis and Promote Nitrogen Metabolism in Medicago truncatula

Bioactive gibberellic acids (GAs) are diterpenoid plant hormones that are biosynthesized through complex pathways and control various aspects of growth and development. Although GA biosynthesis has been intensively studied, the downstream metabolic pathways regulated by GAs have remained largely unexplored. We investigated Tnt1 retrotransposon insertion mutant lines of Medicago truncatula with a dwarf phenotype by forward and reverse genetics screening and phylogenetic, molecular, biochemical, proteomic and metabolomic analyses. Three Tnt1 retrotransposon insertion mutant lines of the gibberellin 3-beta-dioxygenase 1 gene (GA3ox1) with a dwarf phenotype were identified, in which the synthesis of GAs (GA3 and GA4) was inhibited. Phenotypic analysis revealed that plant height, root and petiole length of ga3ox1 mutants were shorter than those of the wild type (Medicago truncatula ecotype R108). Leaf size was also much smaller in ga3ox1 mutants than that in wild-type R108, which is probably due to cell-size diminution instead of a decrease in cell number. Proteomic and metabolomic analyses of ga3ox1/R108 leaves revealed that in the ga3ox1 mutant, flavonoid isoflavonoid biosynthesis was significantly up-regulated, while nitrogen metabolism was down-regulated. Additionally, we further demonstrated that flavonoid and isoflavonoid biosynthesis was induced by prohexadione calcium, an inhibitor of GA3ox enzyme, and inhibited by exogenous GA3. In contrast, nitrogen metabolism was promoted by exogenous GA3 but inhibited by prohexadione calcium. The results of this study further demonstrated that GAs play critical roles in positively regulating nitrogen metabolism and transport and negatively regulating flavonoid biosynthesis through GA-mediated signaling pathways in leaves.

A Retrotransposon Insertion in GhMML3_D12 Is Likely Responsible for the Lintless Locus li3 of Tetraploid Cotton

Cotton (Gossypium) seed fibers can be divided into lint (long) or fuzz (very short). Using fiberless (fuzzless-lintless) mutants, the lint initiation gene Li3 was identified by map-based cloning. The gene is an R2R3-MYB transcription factor located on chromosome D12 (GhMML3_D12). Sequence analysis revealed that li3 is a loss-of-function allele containing a retrotransposon insertion in the second exon that completely blocks the gene’s expression. The genetic loci n2 and n3 underlying the recessive fuzzless phenotype in Gossypium hirsutum were also mapped. The genomic location of n3 overlapped with that of the dominant fuzzless locus N1, and n3 appeared to be a loss-of-function allele caused by a single nucleotide polymorphism (SNP) mutation in the coding region of GhMML3_A12. The n2 allele was found to be co-located with li3 and originated from G. babardense. n2 and li3 are possibly the multiple alleles of the GhMML3_D12 gene. Genetic analysis showed that Li3 and N3 are a pair of homologs with additive effects for the initiation of fibers (fuzz or lint). In addition, the presence of another locus was speculated, and it appeared to show an inhibitory effect on the expression of GhMML3. These findings provide new information about the genetic factors affecting the initiation of fibers in cotton.