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. 2022 Jul 8;13(7):590. doi: 10.1038/s41419-022-05023-0

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© The Author(s) 2022

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 4 — A, B Overexpression or silencing of DCAF1 decreased or increased SAMHD1 expression, respectively. A HL60 cells were transfected with increasing concentrations of HA-tagged DCAF1 24 h, and the levels of the indicated proteins were determined by western blotting. B THP-1 cells were transfected with DCAF1 siRNAs for 24 h, and the levels of the indicated proteins were then determined by western blotting. C DCAF1 interacts with SAMHD1. 293 T cells were transfected with HA-tagged DCAF1 and Myc-tagged SAMHD1 for 24 h. Co-IP was performed with anti-Myc agarose, and the interactions were examined by western blotting. D Overexpression of DCAF1 increased the ubiquitination level of SAMHD1. Myc-tagged SAMHD1 was cotransfected with HA-tagged DCAF1, Flag-ubiquitin or empty vector into 293 T cells for 24 h, and the ubiquitination level of SAMHD1 was determined using IP followed by western blotting. E NONO blocked DCAF1-mediated degradation of SAMHD1. Myc-tagged SAMHD1 and HA-tagged DCAF1 were cotransfected into 293 T cells with or without overexpression of HA-tagged NONO for 24 h. The levels of the indicated proteins were determined by western blotting. F Overexpression of NONO inhibited DCAF1-mediated ubiquitination of SAMHD1. Myc-tagged SAMHD1, HA-tagged DCAF1, and Flag-tagged ubiquitin were cotransfected into 293 T cells with or without overexpression of HA-tagged NONO for 24 h. The ubiquitination of SAMHD1 was evaluated by IP followed by western blotting. G NONO inhibits the interaction of DCAF1 and SAMHD1. Myc-tagged SAMHD1 and HA-tagged DCAF1 were cotransfected into 293 T cells with or without overexpression of Myc-tagged NONO for 24 h. Cells were subsequently subjected to co-IP followed by western blotting with the indicated antibodies. H Silencing of NONO increases the interaction of DCAF1 with SAMHD1. THP-1 cells silencing NONO for 48 h were lysed and were subsequently subjected to co-IP followed by western blotting with the indicated antibodies. For respective immunoblots, the protein levels were quantified by ImageJ software.