Langerhans cell protein 1 (LCP1) binds to PNUTS in the nucleus: implications for this complex in transcriptional regulation

Shin-Jeong Lee,Young-Guen Kwon,Jun-Ki Lee,Young-Myeong Kim,Yong-Sun Maeng

doi:10.3858/emm.2009.41.3.022

Abstract

Protein phosphatase-1 (PP1) nuclear targeting subunit (PNUTS), also called PP1R10, p99, or CAT 53 was originally isolated as a mammalian nuclear PP1-binding protein. In this study, we performed yeast two-hybrid screens to identify PNUTS-interacting proteins. Here, we report that LCP1 (epidermal Langerhans cell protein 1), a novel member of the HMG-box protein family, binds tightly to PNUTS. Co-immunoprecipitation of deletion constructs revealed that the C-terminus of LCP1 is sufficient for the interaction with an N-terminal region of PNUTS that is distinct from its PP1-binding domain. Furthermore, immunofluorescence studies showed that a subpopulation of LCP1 co-localizes with PNUTS in nuclear speckles. Importantly, we found that the N-terminus of LCP1 has a strong trans-activation activity in a GAL4-based heterologous transcription assay. The transcriptional activity of LCP1 is markedly suppressed by its interaction with PNUTS, in a PP1-independent manner. These findings suggest that the coordinated spatial and temporal regulation of LCP1 and PNUTS may be a novel mechanism to control the expression of genes that are critical for certain physiological and pathological processes.

Highlights

Protein phosphatase-1 (PP1) nuclear targeting subunit (PNUTS), known as PP1R10, p99, or CAT 53, was originally isolated as a mammalian nuclear PP1-binding protein (Allen et al, 1998; Kim et al, 2003; Ruma et al, 2005)
We report that Langerhans cell protein 1 (LCP1), which represents a novel member of the HMG-box protein family, is tightly associated with protein phosphatase nuclear targeting subunit (PNUTS) in the mammalian nucleus
We report that LCP1, a mem ber of the TOX HMG-box subfamily, is a novel binding partner of PNUTS in mammalian cells

Summary

Introduction

Protein phosphatase-1 (PP1) nuclear targeting subunit (PNUTS), known as PP1R10, p99, or CAT 53, was originally isolated as a mammalian nuclear PP1-binding protein (Allen et al, 1998; Kim et al, 2003; Ruma et al, 2005). Biochemical analysis has shown that PNUTS binds to PP1 through a consensus PP1-binding 'RVXF motif' (398TVTW401) and to homopolymeric RNA, with high selectivity for poly(A) and poly(G), through a region of its C-terminus that contains RGG motifs (Allen et al, 1998; Kim et al, 2003). We have demonstrated that PNUTS is a hypoxia-inducible gene that regulates the phosphorylation and apoptotic activities of p53 (Lee et al, 2007). Overexpression of PNUTS increases cell death in response to hypoxia through the increased expression of Bax, an apoptosisrelated gene induced by p53 as well as the ubiquitin-dependent proteosomal degradation of MDM2 (Lee et al, 2007).

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