Papaya CpbHLH1/2 regulate carotenoid biosynthesis-related genes during papaya fruit ripening

The ripening of papaya is a physiological and metabolic process associated with accumulation of carotenoids,alternation of flesh color and flavor,which depending on genotype and external factors such as light and hormone.Transcription factors regulating carotenoid biosynthesis have not been analyzed during papaya fruit ripening.RNA-Seq experiments were implemented using different ripening stages of papaya fruit from two papaya varieties.Cis-elements in lycopeneβ-cyclase genes(CpCYC-B and CpLCY-B)were identified,and followed by genome-wide analysis to identify transcription factors binding to these cis-elements,resulting in the identification of CpbHLH1 and CpbHLH2,two bHLH genes.The expressions of CpbHLH1/2 were changed during fruit development,coupled with transcript increase of carotenoid biosynthesis-related genes including CpCYC-B,CpLCY-B,CpPDS2,CpZDS,CpLCY-E,and CpCHY-B.Yeast onehybrid(Y1H)and transient expression assay revealed that CpbHLH1/2 could bind to the promoters of CpCYC-B and CpLCY-B,and regulate their transcriptions.In response to strong light,the results of elevated expression of carotenoid biosynthesis-related genes and the changed expression of CpbHLH1/2 indicated that CpbHLH1/2 were involved in light-mediated mechanisms of regulating critical genes in the carotenoid biosynthesis pathway.Collectively,our findings demonstrated several TF family members participating in the regulation of carotenoid genes and proved that CpbHLH1 and CpbHLH2 individually regulated the transcription of lycopeneβ-cyclase genes(CpCYC-B and CpLCY-B).This study yielded novel findings on regulatory mechanism of carotenoid biosynthesis during papaya fruit ripening.

Advances in NK cell production

Immunotherapy based on natural killer(NK)cells is a promising approach for treating a variety of cancers.Unlike T cells,NK cells recognize target cells via a major histocompatibility complex(MHC)-independent mechanism and,without being sensitized,kill the cells directly.Several strategies for obtaining large quantities of NK cells with high purity and high cytotoxicity have been developed.These strategies include the use of cytokine−antibody fusions,feeder cells or membrane particles to stimulate the proliferation of NK cells and enhance their cytotoxicity.Various materials,including peripheral blood mononuclear cells(PBMCs),umbilical cord blood(UCB),induced pluripotent stem cells(iPSCs)and NK cell lines,have been used as sources to generate NK cells for immunotherapy.Moreover,genetic modification technologies to improve the proliferation of NK cells have also been developed to enhance the functions of NK cells.Here,we summarize the recent advances in expansion strategies with or without genetic manipulation of NK cells derived from various cellular sources.We also discuss the closed,automated and GMP-controlled large-scale expansion systems used for NK cells and possible future NK cell-based immunotherapy products.