Nuclear protein IK undergoes dynamic subcellular translocation and forms unique nuclear bodies during the cell cycle
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RESEARCH
Open Access
Nuclear protein IK undergoes dynamic subcellular translocation and forms unique nuclear bodies during the cell cycle Liyan Hu1, Feikun Yang1, Xianan Liu1, Dazhong Xu1 and Wei Dai1,2*
Abstract IK is a nuclear protein containing a unique domain named RED due to the presence of a repetitive arginine (R), aspartic (E), and glutamic acid (D) sequence. To date, the function of this protein remains largely unknown despite of a couple of previous studies in the literature. Here we report that depletion of IK via RNA interference results in mitotic arrest. We also demonstrate that IK undergoes dynamic translocation during interphase and mitosis. In particular, IK is primarily present in some interphase cells as nuclear foci/bodies which do not co-localize with nucleoli, PMA bodies and Cajal bodies. Pull-down analysis coupled with mass spectrometry reveals that IK is associated with DHX15, a putative ATP-dependent RNA helicase. Our results strongly suggest that IK may participate in pre-mRNA splicing and that it may be a useful biomarker for a new nuclear structure in the cell.
Introduction IK was originally identified as a cytokine that inhibits interferon gamma (IFN-γ)-induced expression of HLA class II antigen during immune responses in K562 erythroleukemic cell line [1]. The protein got its name IK based on these findings. Human IK contains 557 amino acids and migrates at about 80 kDa on SDS-PAGE [2]. It is also named RED owing to the presence of a repetitive arginine (R), aspartic (E), and glutamic acid (D) sequence [2]. It has also been reported that IK is one of the spliceosome factors [3,4]. Screening of an siRNA library containing 23,835 human genes reveals that depletion of IK induces mitotic arrest, primarily characterized by having a high mitotic index [5]. A recent study shows that IK is required for the localization of MAD1, a spindle checkpoint protein, to the kinetochores and involved in the regulation of the spindle assembly checkpoint [6]. In the present study, we have confirmed that depletion of IK causes mitotic arrest. Our further investigation reveals that the subcellular localization of IK is dynamic during the cell cycle. We also show that the expression of IK is cell cycle-regulated. Affinity pull-down and mass * Correspondence: [email protected] 1 Department of Environmental Medicine, New York University Langone Medical Center, Tuxedo, NY 10987, USA 2 Department of Biochemistry & Molecular Pharmacology, New York University Langone Medical Center, Tuxedo, NY 10987, USA
spectrometry analyses reveal that IK interacts with DHX15, a putative ATP-dependent RNA helicase which is implicated in pre-mRNA splicing. Our current study suggests that IK can be explored as a new biomarker for cell proliferation and checkpoint control. Materials and methods Cell culture
HeLa cell line was originally obtained from the American Type Culture Collection (Manassas, VA). Cells were cultured in DMEM supplemented with 10% fetal bovine serum (FBS, Invitrogen, Carlsbad, CA) and antibiotics (100 μg/m
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