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Cell type-dependent suppression of the RNA helicase DDX3Y levels by the close paralog DDX3X
Xiaolu Xu1, Jessica Rainey1, Nicholas F Grigoropoulos2,3
1Department of Biological Sciences, University of Delaware, Newark, DE, United States.
Abstract:
The DEAD-box RNA helicase DDX3X has important roles in development and disease. Loss of DDX3X during developmental and pathological processes such as tumorigenesis can lead to compensatory upregulation of the close paralog DDX3Y in males, which may underlie the sexual dimorphism displayed by some DDX3X-associated diseases. However, how DDX3X cross-regulates DDX3Y remains largely unknown. Here, we investigated the regulation of DDX3Y by DDX3X in two male-derived human cancer cell lines, HCT116 and U87MG. Depletion of DDX3X in HCT116 cells results in moderately increased DDX3Y mRNA and protein, in part due to stabilization of DDX3Y transcripts. Conversely, reduction of DDX3X in U87MG cells markedly upregulates DDX3Y protein without affecting its mRNA, mainly by enhancing DDX3Y protein stability. We further show that DDX3X physically interacts with DDX3Y. DDX3Y is much less stable than DDX3X in U87MG cells, and substitution of two lysine residues in DDX3Y with the corresponding arginine in DDX3X stabilizes DDX3Y. Thus, the compensatory upregulation of DDX3Y following DDX3X loss can occur at either transcript or protein level, suggesting complex and cell type-specific cross-regulation between these X- and Y-linked paralogs to keep the total DDX3 dosage in check.
Insights
Loss of DDX3X RNA helicase can cause compensatory DDX3Y upregulation in males. This study reveals DDX3X regulates DDX3Y at transcript or protein levels, depending on cell type, to maintain DDX3 dosage.
Area of Science:
- Molecular Biology
- Genetics
- Cancer Research
Background:
- The DEAD-box RNA helicase DDX3X is crucial in development and disease.
- Loss of DDX3X can lead to DDX3Y upregulation in males, potentially explaining sexual dimorphism in DDX3X-related diseases.
- Mechanisms of DDX3X-DDX3Y cross-regulation are poorly understood.
Purpose of the Study:
- To investigate how DDX3X regulates its paralog DDX3Y in male human cancer cell lines.
- To elucidate the molecular mechanisms underlying DDX3Y upregulation upon DDX3X loss.
Main Methods:
- Depletion of DDX3X in HCT116 and U87MG cell lines.
- Analysis of DDX3Y mRNA and protein levels.
- Investigation of transcript and protein stability.
- Assessment of physical interaction between DDX3X and DDX3Y.
- Site-directed mutagenesis to study protein stability.
Main Results:
- DDX3X depletion moderately increased DDX3Y mRNA and protein in HCT116 cells, partly via transcript stabilization.
- DDX3X reduction markedly upregulated DDX3Y protein without altering mRNA in U87MG cells, mainly through enhanced protein stability.
- DDX3X physically interacts with DDX3Y.
- Specific lysine residues in DDX3Y influence its stability, with substitution mimicking DDX3X conferring greater stability.
Conclusions:
- DDX3X-mediated regulation of DDX3Y occurs through distinct mechanisms (transcript vs. protein stability) in a cell type-specific manner.
- These findings highlight complex cross-regulation between X- and Y-linked DDX3 paralogs to maintain total DDX3 dosage.
- Understanding this regulation is key to addressing sexual dimorphism in DDX3X-associated diseases.
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