Integral UBL domain proteins: a family of proteasome interacting proteins

Abstract

The family of ubiquitin-like (UBL) domain proteins (UDPs) comprises a conserved group of proteins involved in a multitude of different cellular activities. However, recent studies on UBL-domain proteins indicate that these proteins appear to share a common property in their ability to interact with 26S proteasomes.

The 26S proteasome is a multisubunit protease which is responsible for the majority of intracellular proteolysis in eukaryotic cells. Before degradation commences most proteins are first marked for destruction by being coupled to a chain of ubiquitin molecules. Some UBL-domain proteins catalyse the formation of ubiquitin–protein conjugates, whereas others appear to target ubiquitinated proteins for degradation and interact with chaperones. Hence, by binding to the 26S proteasome the UBL-domain proteins seem to tailor and direct the basic proteolytic functions of the particle to accommodate various cellular substrates.

Introduction

The degradation of proteins in eukaryotic cells plays a pivotal role in the regulation of intracellular levels of key proteins involved in a series of cellular functions including metabolism, cell cycle regulation and gene expression [1]. In higher eukaryotes, a continuous protein degradation system is also critical for immunological reactions through the formation of antigenic peptides for presentation on the major histocompatibility complex (MHC) class I [2].

Before they are degraded, most proteins are covalently conjugated to ubiquitin by an enzymatic cascade involving three enzymes, called E1, E2 and E3 [3], [4]. Eukaryotic cells contain only one E1 enzyme, a few E2 enzymes and many E3 enzymes. First ubiquitin is bound to a ubiquitin activating enzyme, E1, in an ATP-consuming process. Subsequently, the activated ubiquitin moiety is transferred to a ubiquitin conjugating enzyme, E2. Ubiquitin-loaded E2s then associate with substrate-specific ubiquitin–protein ligases, E3s, which catalyse the formation of an isopeptide bond between the C terminus of ubiquitin and an amino group, typically on a lysine residue, either directly on the target protein or on the last ubiquitin moiety of a ubiquitin chain attached to the target protein. Finally, some proteins require the action of a ubiquitin chain elongation factor, E4, for efficient multiubiquitination [5]. Several such rounds of ubiquitination yields proteins carrying chains of ubiquitin moieties. However, the process of ubiquitination is reversible and several deubiquitinating enzymes play important regulatory roles in trimming or cleaving the ubiquitin chains, prior to degradation [6].

Once the ubiquitin chain on a target protein becomes at least four moieties long, it facilitates the recognition of the target protein by the 26S proteasome [7], a 2.5 MDa multisubunit proteolytic particle which resides in the nucleus and cytoplasm of all eukaryotic cells [8].

Structural exploration of the 26S proteasome by various experimental approaches has revealed that the 26S proteasome is composed of two subcomplexes, a 20S core particle and a 19S regulatory complex [8].

The proteolytic activity resides within the 20S core complex which forms a cylindrical structure composed of four stacked heptameric rings housing a central chamber flanked by two antechambers [9]. The catalytic sites face the interior of the central chamber that is only accessible through narrow pores at either end of the 20S cylinder [9]. Substrate entry through these pores is guarded by the 19S regulatory complexes which associate with the ends of the 20S cylinder [10].

Tightly regulating access to the active sites of the 26S proteasome obviously prevents the hazard of indiscriminate proteolysis within the cell, and as generally only unfolded polypeptides may fit through the narrow entry pores, one function of the 19S particle is to provide a means to unfold the substrate prior to degradation [11], [12].

The 19S particle can be dissociated into two subcomplexes called the base and the lid [13]. The base subcomplex is located proximally to the 20S proteasome and contains six AAA (ATPases associated with various cellular activities) family ATPase subunits, which are believed to mediate the substrate-unfolding step alluded to above. Additionally, the base contains two large non-ATPase subunits called Rpn1 and Rpn2, and as a multitude of different proteins have been found to interact with these subunits [14], they appear to play a central role in providing a scaffold or platform for transiently associated proteins.

Relative to the 20S core particle the more distally located lid subcomplex covers the base and is thought to be involved in the deubiquitination of substrates prior to their translocation and degradation [15], [16].