DOR/Tp53inp2 and Tp53inp1 Constitute a Metazoan Gene Family Encoding Dual Regulators of Autophagy and Transcription

Ana Sancho,Javier U Chicote,Antonio García-España,Antonio Zorzano,Caroline Mauvezin,Miriam Royo,Rob Desalle,Jordi Duran,Trond Lamark,Maria J Macias,David Sala,Terje Johansen,Manuel Palacín,Endalkachew A Alemu

doi:10.1371/journal.pone.0034034

Abstract

Human DOR/TP53INP2 displays a unique bifunctional role as a modulator of autophagy and gene transcription. However, the domains or regions of DOR that participate in those functions have not been identified. Here we have performed structure/function analyses of DOR guided by identification of conserved regions in the DOR gene family by phylogenetic reconstructions. We show that DOR is present in metazoan species. Invertebrates harbor only one gene, DOR/Tp53inp2, and in the common ancestor of vertebrates Tp53inp1 may have arisen by gene duplication. In keeping with these data, we show that human TP53INP1 regulates autophagy and that different DOR/TP53INP2 and TP53INP1 proteins display transcriptional activity. The use of molecular evolutionary information has been instrumental to determine the regions that participate in DOR functions. DOR and TP53INP1 proteins share two highly conserved regions (region 1, aa residues 28–42; region 2, 66–112 in human DOR). Mutation of conserved hydrophobic residues in region 1 of DOR (that are part of a nuclear export signal, NES) reduces transcriptional activity, and blocks nuclear exit and autophagic activity under autophagy-activated conditions. We also identify a functional and conserved LC3-interacting motif (LIR) in region 1 of DOR and TP53INP1 proteins. Mutation of conserved acidic residues in region 2 of DOR reduces transcriptional activity, impairs nuclear exit in response to autophagy activation, and disrupts autophagy. Taken together, our data reveal DOR and TP53INP1 as dual regulators of transcription and autophagy, and identify two conserved regions in the DOR family that concentrate multiple functions crucial for autophagy and transcription.

Highlights

Macroautophagy is a major cellular pathway for the degradation of long-lived proteins and organelles [1,2]
Identification of the DOR/Tp53inp2 gene family To retrieve as many sequences belonging to the DOR gene family as possible, we performed blast searches using known sequences of DOR/TP53INP2 and TP53INP1 (TP53INP1 is a DOR homologous protein named SIP, Teap or Stinp, [14,24,25,26] in the NCBI ESTs and protein databases)
DOR regions 1 and 2 contain signals that are crucial for autophagic activity we studied whether regions 1 and 2 in the DOR sequence contribute to the autophagic activity exerted by this protein in mammalian cells [12,13] and in fruit flies [12]

Summary

Introduction

Macroautophagy (here referred to as autophagy) is a major cellular pathway for the degradation of long-lived proteins and organelles [1,2]. Our understanding of autophagy has increased substantially in recent years, there are still many open questions related to the identity of the components of the autophagic pathway, the molecular signals that regulate it, and the mechanisms that target specific cellular components to be degraded. In this respect, there is evidence of a bidirectional functional link between nuclear regulators of gene transcription and autophagy. Some autophagic proteins, such as Beclin 1, p62 or LC3, undergo nucleocytoplasmic shuttling in cells [9,10,11], which may be relevant to the regulation of autophagy

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Results

Conclusion