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The piRNA amplification pathway in Bombyx is operated by Ago3 and Siwi in their piRISC form. The DEAD‐box protein, Vasa, facilitates Ago3‐piRISC production by liberating cleaved RNAs from Siwi‐piRISC in an ATP hydrolysis‐dependent manner. However, the Vasa‐like factor facilitating Siwi‐piRISC production along this pathway remains unknown. Here, we identify DEAD‐box polypeptide 43 (DDX43) as the Vasa‐like protein functioning in Siwi‐piRISC production. DDX43 belongs to the helicase superfamily II along with Vasa, and it contains a similar helicase core. DDX43 also contains a K‐homology (KH) domain, a prevalent RNA‐binding domain, within its N‐terminal region. Biochemical analyses show that the helicase core is responsible for Ago3‐piRISC interaction and ATP hydrolysis, while the KH domain enhances the ATPase activity of the helicase core. This enhancement is independent of the RNA‐binding activity of the KH domain. For maximal DDX43 RNA‐binding activity, both the KH domain and helicase core are required. This study not only provides new insight into the piRNA amplification mechanism but also reveals unique collaborations between the two domains supporting DDX43 function within the pathway. Synopsis: Vasa facilitates Ago3‐piRISC production in the ping‐pong cycle by liberating cleaved RNAs from Siwi‐piRISC. This study shows that DDX43 is Vasa's counterpart facilitating Siwi‐piRISC production by liberating cleaved RNAs from Ago3‐piRISC. The DEAD‐box protein DDX43 interconnects Ago3‐piRISC with unbound Siwi in the ping‐pong cycle.DDX43 facilitates Siwi‐piRISC production by liberating cleaved RNAs from Ago3‐piRISC.The helicase core of DDX43 is responsible for DDX43‐Ago3 interaction and ATP hydrolysis.The KH domain of DDX43 enhances the ATPase activity of the helicase core independently of its own RNA‐binding activity. [ABSTRACT FROM AUTHOR]

The clinical activity of decitabine (5‐aza‐2‐deoxycytidine, DAC), a hypomethylating agent, has been demonstrated in acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS) patients. However, secondary resistance to this agent often occurs during treatment and leads to treatment failure. It is important to clarify the mechanisms underlying the resistance for improving the efficacy. In this study, by gradually increasing concentration after a continuous induction of DAC, we established the DAC‐resistant K562 cell line (K562/DAC) from its parental cell line K562. The proliferation and survival rate of K562/DAC was significantly increased, whereas the apoptosis rate was remarkably decreased than that of K562 after DAC treatment. In K562/DAC, a total of 108 genes were upregulated and 118 genes were downregulated by RNA‐Seq. In addition, we also observed aberrant expression of DDX43/H19/miR‐186 axis (increased DDX43/H19 and decreased miR‐186) in K562/DAC cells. Ectopic expression of DDX43 in parental K562 cells rendered cells resistant to the DAC. Taken together, we successfully established DAC‐resistant K562 cell line which can serve as a good model for investigating DAC resistance mechanisms, and DDX43/H19/miR‐186 may be involved in DAC resistance in K562. [ABSTRACT FROM AUTHOR]

This paper aims to determine the expression and clinical significance of DDX43 in lung adenocarcinoma. Expression of DDX43 gene and protein of lung adenocarcinoma tissue and para-carcinoma tissues was observed in 27 cases by RT-PCR and immunohistochemistry. These patients were diagnosed as lung adenocarcinoma in the Huaihe Hospital of Henan University from February 2015 to December 2015. The relative ratio of DDX43 mRNA expression in lung adenocarcinoma and para-carcinoma tissues was 0.87±0.62 versus 0.21±0.77 and the difference between the two groups was statistically significant (P<0.01). The expression of DDX43 in normal lung tissues and lung adenocarcinoma tissues was different. The positive rate of DDX43 expression in lung adenocarcinoma tissues was significantly higher than that in normal lung tissues, and the difference was statistically significant (P<0.05). The analysis of clinical pathological characteristics showed that the increase of protein expression was related to the stage and metastasis of lung adenocarcinoma. DDX43 is highly expressed in lung adenocarcinoma, and the expression level is related to the stage and metastasis of lung adenocarcinoma, suggesting that DDX43 is closely related to the occurrence and development of lung adenocarcinoma, and may be a molecular marker for early diagnosis of lung adenocarcinoma. [ABSTRACT FROM AUTHOR]

The K-homology (KH) domain is a nucleic acid-binding domain present in many proteins. Recently, we found that the DEAD-box helicase DDX43 contains a KH domain in its N-terminus; however, its function remains unknown. Here, we purified recombinant DDX43 KH domain protein and found that it prefers binding ssDNA and ssRNA. Electrophoretic mobility shift assay and NMR revealed that the KH domain favors pyrimidines over purines. Mutational analysis showed that the GXXG loop in the KH domain is involved in pyrimidine binding. Moreover, we found that an alanine residue adjacent to the GXXG loop is critical for binding. Systematic evolution of ligands by exponential enrichment, chromatin immunoprecipitation-seq, and cross-linking immunoprecipitation-seq showed that the KH domain binds C-/T-rich DNA and U-rich RNA. Bioinformatics analysis suggested that the KH domain prefers to bind promoters. Using 15 N-heteronuclear single quantum coherence NMR, the optimal binding sequence was identified as TTGT. Finally, we found that the full-length DDX43 helicase prefers DNA or RNA substrates with TTGT or UUGU single-stranded tails and that the KH domain is critically important for sequence specificity and unwinding processivity. Collectively, our results demonstrated that the KH domain facilitates the substrate specificity and processivity of the DDX43 helicase., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2020 The Authors. Published by Elsevier Inc. All rights reserved.)