Mechanisms of Small Heat Shock Proteins

Abstract

Small heat shock proteins (sHSPs) are ATP-independent chaperones that delay formation of harmful protein aggregates. sHSPs' role in protein homeostasis has been appreciated for decades, but their mechanisms of action remain poorly understood. This gap in understanding is largely a consequence of sHSP properties that make them recalcitrant to detailed study. Multiple stress-associated conditions including pH acidosis, oxidation, and unusual availability of metal ions, as well as reversible stress-induced phosphorylation can modulate sHSP chaperone activity. Investigations of sHSPs reveal that sHSPs can engage in transient or long-lived interactions with client proteins depending on solution conditions and sHSP or client identity. Recent advances in the field highlight both the diversity of function within the sHSP family and the exquisite sensitivity of individual sHSPs to cellular and experimental conditions. Here, we will present and highlight current understanding, recent progress, and future challenges.

Figure 1.

General structural features of small heat shock proteins (sHSPs). (A) Sequence alignment of the 10 human sHSPs and Sip 1, a pH-sensitive sHSP from Caenorhabditis elegans generated with clustal omega and visualized with Jalview. The colored bar above the sequences identify the three regions of sHSPs: amino-terminal region (NTR) (blue), α-crystallin domain (ACD) (gray), and carboxy-terminal region (CTR) (red). Important sequence elements are highlighted in different colors: the conserved amino-terminal sequence (yellow), the β4 and β8 strands that compose the groove (gray), the β6 + 7 strand that makes the dimer interface (green), and amino- and carboxy-terminal I/V-X-I/V motifs (blue). Histidine residues are highlighted in orange. The positions with sidechains pointing into the β4/β8 groove are indicated by purple dots, and the site of the “bump mutation” located in β8 is additionally labeled by a green bar crossing the purple dot. (B) ACD architecture. All sHSPs contain a core conserved ACD with an IgG-like β-sandwich fold. Human ACDs form an antiparallel dimer along the β6 + 7 strand. (C) Oligomeric organization of HSPB5. Two pseudo-atomic models of HSPB5 oligomers generated using a combination of solid-state nuclear magnetic resonanace (NMR), electron microscopy (EM), small-angle X-ray scattering, and structural modeling (left) (Jehle et al. 2011) and NMR, EM, and structural modeling (right) (Braun et al. 2011). Both depict 24-mers with tetrahedral geometry and extensive interactions among the three regions.

Figure 2.

Structural aspects of small heat shock protein (sHSP) assembly. Assembly of sHSP oligomers is driven by interactions that involve all three regions: the α-crystallin domain (ACD) (gray), the amino-terminal region (NTR) (blue), and the carboxy-terminal region (CTR) (red). PDB IDs are indicated below each panel. (A) ACD β4/β8 groove interactions with carboxy-terminal I/V-X-I/V motifs. The CTRs of many human sHSPs contain I/V-X-I/V motifs, which can dock into the β4/β8 groove on the outer edge of the ACD. Shown here, HSPB1 and HSPB5 ACDs were crystallized with peptides containing their carboxy-terminal I/V-X-I/V motifs (Hochberg et al. 2014). (B) ACD β4/β8 groove interactions with an amino-terminal I/V-X-I/V motif. Several human sHSPs contain I/V-X-I/V motifs in their NTRs, which can also bind the β4/β8 groove. A crystal structure of full-length HSPB6 revealed its amino-terminal VPV motif bound in the groove (Sluchanko et al. 2017). (C) ACD dimer interface interactions with an NTR sequence. The HSPB6 structure revealed interactions between the conserved NTR sequence ²⁷RLFDQ³¹ and a groove at the dimer interface of the ACD (Sluchanko et al. 2017). (D) Phosphorylation-dependent NTR–client interactions. Phosphorylated Ser16 (teal) in the HSPB6 NTR facilitates interactions with the client protein 14-3-3 (orange). This may serve as a model for other phosphorylation-dependent client interactions and provides the only atomic-level information about NTR–client interactions available to date (Sluchanko et al. 2017).

Figure 3.

Diversity of small heat shock protein (sHSP) chaperone mechanisms. Many human sHSPs form polydisperse oligomers whose average number of subunits is sensitive to solution conditions or posttranslational modifications. Client aggregation is also sensitive to solution conditions and can potentially affect the client states recognized by an sHSP. It is unknown whether all subunits within a given sHSP ensemble are chaperone active. Many open questions remain, including: (1) Which client states and features of clients do sHSPs recognize? (2) Do different sHSP ensemble sizes recognize different client states and therefore have different chaperone activity? The answers to these and other questions likely depend on sHSPs and client identities and the solution conditions used to assess chaperone function.