Werner complex deficiency in cells disrupts the Nuclear Pore Complex and the distribution of lamin B1.
From the surrounding shell to the inner machinery, nuclear proteins provide the functional plasticity of the nucleus. This study highlights the nuclear association of Pore membrane (POM) protein NDC1 and Werner protein (WRN), a RecQ helicase responsible for the DNA instability progeria disorder, Werner Syndrome. In our previous publication, we connected the DNA damage sensor Werner's Helicase Interacting Protein (WHIP), a binding partner of WRN, to the NPC. Here, we confirm the association of the WRN/WHIP complex and NDC1. In established WRN/WHIP knockout cell lines, we further demonstrate the interdependence of WRN/WHIP and Nucleoporins (Nups). These changes do not completely abrogate the barrier of the Nuclear Envelope (NE) but do affect the distribution of FG Nups and the RAN gradient, which are necessary for nuclear transport. Evidence from WRN/WHIP knockout cell lines demonstrates changes in the processing and nucleolar localization of lamin B1. The appearance of "RAN holes" void of RAN corresponds to regions within the nucleolus filled with condensed pools of lamin B1. From WRN/WHIP knockout cell line extracts, we found three forms of lamin B1 that correspond to mature holoprotein and two potential post-translationally modified forms of the protein. Upon treatment with topoisomerase inhibitors lamin B1 cleavage occurs only in WRN/WHIP knockout cells. Our data suggest the link of the NDC1 and WRN as one facet of the network between the nuclear periphery and genome stability. Loss of WRN complex leads to multiple alterations at the NPC and the nucleolus.
Department of Chemistry, Chemical Biology & Biomedical Engineering, Stevens Institute of Technology, Hoboken, NJ 07030, USA.
Zhi
Z
Li
Department of Chemistry, Chemical Biology & Biomedical Engineering, Stevens Institute of Technology, Hoboken, NJ 07030, USA.
Yizhou
Y
Zhu
Department of Chemistry, Chemical Biology & Biomedical Engineering, Stevens Institute of Technology, Hoboken, NJ 07030, USA.
Yujia
Y
Zhai
Department of Chemistry, Chemical Biology & Biomedical Engineering, Stevens Institute of Technology, Hoboken, NJ 07030, USA.
Michelle
M
R Castroagudin
Department of Chemistry, Chemical Biology & Biomedical Engineering, Stevens Institute of Technology, Hoboken, NJ 07030, USA.
Yifei
Y
Bao
Department of Chemistry, Chemical Biology & Biomedical Engineering, Stevens Institute of Technology, Hoboken, NJ 07030, USA.
Tommy E
TE
White
Department of Chemistry, Chemical Biology & Biomedical Engineering, Stevens Institute of Technology, Hoboken, NJ 07030, USA. Electronic address: jglavy@stevens.edu.
Joseph S
JS
Glavy
D000818:Animals; D015153:Blotting, Western; D002645:Chickens; D004249:DNA Damage; D004260:DNA Repair; D004268:DNA-Binding Proteins / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D055786:Gene Knockout Techniques; D034921:Lamin Type B / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism; D022022:Nuclear Pore / Q000378:metabolism*; D028861:Nuclear Pore Complex Proteins / Q000378:metabolism; D011485:Protein Binding; D014898:Werner Syndrome / Q000378:metabolism*
D004268:DNA-Binding Proteins; D034921:Lamin Type B; D008565:Membrane Proteins; D028861:Nuclear Pore Complex Proteins; C093413:lamina-associated polypeptide 2
CPT; Camptothecin; Etop; Etoposide; HU; Hydroxyurea; IP; Lamin B1; Lamina-associated polypeptide 2; Lap2β; NDC1; NE; NPC; Nuclear Envelope; Nuclear Pore Complex; Nucleolus; Nucleoporins; Nups; POM; Pore membrane proteins; WHIP; WRN; Werner; Werner protein; Werner's Helicase Interacting Protein; immunoprecipitation