Expansin SlExp1 and endoglucanase SlCel2 synergistically promote fruit softening and cell wall disassembly in tomato.

Fruit softening, an irreversible process that occurs during fruit ripening, can lead to losses and waste during postharvest transportation and storage. Cell wall disassembly is the main factor leading to loss of fruit firmness, and several ripening-associated cell wall genes have been targeted for genetic modification, particularly pectin modifiers. However, individual knockdown of most cell wall-related genes has had minimal influence on cell wall integrity and fruit firmness, with the notable exception of pectate lyase. Compared to pectin disassembly, studies of the cell wall matrix, the xyloglucan-cellulose framework, and underlying mechanisms during fruit softening are limited. Here, a tomato (Solanum lycopersicum) fruit ripening-associated α-expansin (SlExpansin1/SlExp1) and an endoglucanase (SlCellulase2/SlCel2), which function in the cell wall matrix, were knocked out individually and together using clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated nuclease 9-mediated genome editing. Simultaneous knockout of SlExp1 and SlCel2 enhanced fruit firmness, reduced depolymerization of homogalacturonan-type pectin and xyloglucan, and increased cell adhesion. In contrast, single knockouts of either SlExp1 or SlCel2 did not substantially change fruit firmness, while simultaneous overexpression of SlExp1 and SlCel2 promoted early fruit softening. Collectively, our results demonstrate that SlExp1 and SlCel2 synergistically regulate cell wall disassembly and fruit softening in tomato.

FBXO30 functions as a tumor suppressor and an E3 ubiquitin ligase for hZIP1­mediated HIF­1α degradation in renal cell carcinoma.

Studies on clear cell renal cell carcinoma (ccRCC) are gaining momentum due to its high malignancy and potential to metastasize. F­box protein 30 (FBXO30) is a member of the F­box protein family; however, its role and mechanism in cancer remains to be fully elucidated. Western blotting, reverse transcription­quantitative PCR and immunohistochemsitry were performed to detect the expression levels of FBXO30 in ccRCC tissues and adjacent normal tissues. Tumor biological function assays and animal experiments were conducted to clarify the inhibitory effect of FBXO30 on the progression and metastasis of ccRCC. Protein half­life assay, MG132 inhibition assay, immunofluorescence assay and co­immunoprecipitation assay were performed to explore the ubiquitination mechanism of FBXO30 and HIF­1α. Zinc supplementation assay was used to verify the regulatory relationship between human ZRT, IRT­like protein 1 (hZIP1), FBXO30 and HIF­1α. The present study revealed that the expression levels of FBXO30 were lower in ccRCC tissues compared with those in normal adjacent tissues. In addition, FBXO30 inhibited the tumorigenesis and metastatic capacity of ccRCC cells in vivo and in vitro. FBXO30 mediated the ubiquitination and degradation of hypoxia­inducible factor­1α (HIF­1α) in ccRCC cells under normoxia, thereby inhibiting the oncogenic effect of HIF­1α. Notably, hZIP1 served as an upstream regulator of FBXO30, regulating the expression of FBXO30 and HIF­1α by recruiting Zn2+. In conclusion, the present data suggested that FBXO30 is a novel E3 ubiquitination ligase that can function as a tumor suppressor in ccRCC, and the hZIP1/Zn2+/FBXO30/HIF­1α axis may provide potential biomarkers or therapeutic targets for ccRCC.

An EjbHLH14-EjHB1-EjPRX12 module is involved in methyl jasmonate alleviation of chilling-induced lignin deposition in loquat fruit.

Loquat fruit are susceptible to chilling injuries induced by postharvest storage at low temperature. The major symptoms are increased lignin content and flesh firmness, which cause a leathery texture. Pretreatment with methyl jasmonate (MeJA) can alleviate this low-temperature-induced lignification, but the mechanism is not understood. In this study, we characterized a novel class III peroxidase, EjPRX12, and studied its relationship to lignification. Transcript levels of EjPRX12 were attenuated following MeJA pretreatment, consistent with the reduced lignin content in fruit. In vitro enzyme activity assay indicated that EjPRX12 polymerized sinapyl alcohol, and overexpression of EjPRX12 in Arabidopsis promoted lignin accumulation, indicating that it plays a functional role in lignin polymerization. We also identified an HD-ZIP transcription factor, EjHB1, repressed by MeJA pretreatment, which directly bound to and significantly activated the EjPRX12 promoter. Overexpression of EjHB1 in Arabidopsis promoted lignin accumulation with induced expression of lignin-related genes, especially AtPRX64. Furthermore, a JAZ-interacting repressor, EjbHLH14, was characterized, and it is proposed that MeJA pretreatment caused EjbHLH14 to be released to repress the expression of EjHB1. These results identified a novel regulatory pathway involving EjbHLH14-EjHB1-EjPRX12 and revealed the molecular mechanism whereby MeJA alleviated lignification of loquat fruit at low temperature.