overexpression of h-THOR in zebrafish was sufficient for a significant increase in not only the onset of melanoma development , but also on the size of the melanoma tumors that developed
This method provided a powerful way to identify the THOR-IGF2BP1 interaction as one conserved across multiple vertebrate species The binding of lncRNAs to RNA-binding proteins (RBPs) has been reported to be widespread (Li et al., 2015), and multiple efforts have been undertaken to characterize the functional implication of lncRNA-RBP interaction beyond promiscuous binding
6、Human THOR Enhances the Onset of Melanoma in Zebrafish
p53 knockout (p53-/-) zebrafish to enhance the melanoma phenotype observed as previously described
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zebrafish animal model
z-THOR-knockout zebrafish line
zebrafish produced a significant fertilization defect in the zebrafish offspring, supporting the role of THOR in the testis and suggesting a primary functional role for THOR in fertility
THORoncogenic long non-coding RNA)
汇报者： 导 师： 2018年7月
IGF2 mRNA-binding proteins (IGF2BPs)
Result
1、Discovery of THOR, a Conserved lncRNA Expressed in the Testis Large-scale transcriptome sequence
3、Characterization of the THOR-IGF2BP1 interaction
zebrafish THOR added to human H1299 cell lysate (green)
human THOR added to human H1299 cell lysate (blue) zebrafish THOR added to zebrafish embryo lysate (yellow) human THOR added to zebrafish embryo lysate (red)
THOR knockdown reduced IGF2BP1 binding of both IGF2 and CD44 while overexpression of THOR increased IGF2 and CD44 binding
5、THOR-Knockout Zebrafish Exhibit Fertilization Defects and Resistance to Melanoma Formation • THOR: Sequence conservation A conserved tissue expression pattern Conservation of its binding interaction to IGF2BP1 The potential function of THOR in a different animal model
Additionally, to further interrogate in vivo function of THOR, we produced a zebrafish melanoma model that employs embryonic injection of human NRAS-K61 driven by the mitfa promoter (a zebrafish gene expressed in melanocytes), resulting in the formation of easily visible zebrafish melanoma The fertilization defect and resistance to melanoma mediated by THOR present compelling evidence for a conserved role of THOR in vertebrate physiology and pathophysiology
IGF2BP1, in conjunction with number of other proteins comprising the messenger ribonucleoprotein (mRNP) complex, has been implicated in mediating RNA stability and translation through its binding to a number of well-defined mRNA targets
Direct binding interaction of THOR and IGF2BP1 was confirmed via addition of in vitro transcribed h-THOR to purified myc-tagged IGF2BP1
4、THOR Regulates IGF2BP1’s Target mRNAs Levels
(Genotype-Tissue Expression)
qRT-PCR
ISH and H&E
2、Expression and Functional Implication of THOR in Human Cancers
lung squamous carcinoma (LUSC) lung adenocarcinoma (LUAD) H1299: human lung adenocarcinoma cell line