Dynamic Subunit Exchange and the Regulation of Microtubule Assembly by the Stress Response Protein Human αB Crystallin

Abstract

BackgroundThe small heat shock protein (sHSP), human αB crystallin, forms large, polydisperse complexes that modulate the tubulin-microtubule equilibrium using a dynamic mechanism that is poorly understood. The interactive sequences in αB crystallin for tubulin are surface exposed, and correspond to interactive sites for the formation of αB crystallin complexes.Methodology/Principal FindingsThere is sequence homology between tubulin and the interactive domains in the β8-strand of the core domain and the C-terminal extension of αB crystallin. This study investigated the hypothesis that the formation of tubulin and αB crystallin quaternary structures was regulated through common interactive domains that alter the dynamics of their assembly. Size exclusion chromatography (SEC), SDS-PAGE, microtubule assembly assays, aggregation assays, multiple sequence alignment, and molecular modeling characterized the dynamic response of tubulin assembly to increasing concentrations of αB crystallin. Low molar ratios of αB crystallin∶tubulin were favorable for microtubule assembly and high molar ratios of αB crystallin∶tubulin were unfavorable for microtubule assembly. Interactions between αB crystallin and unassembled tubulin were observed using SEC and SDS-PAGE.Conclusions/SignificanceSubunits of αB crystallin that exchange dynamically with the αB crystallin complex can interact with tubulin subunits to regulate the equilibrium between tubulin and microtubules.

Proteomic changes to the sarcoplasmic fraction of predominantly red or white muscle following acute heat stress

“…In addition to typical Hsp chaperone and anti-apoptotic functions, α-crystallin B chain may also play a role in desmin and other intermediate filament assembly, reactive oxygen species regulation, and microfilament assembly [40][41][42][43]. Because the antibody used in this study did not detect these two spots, further LC-MS/MS was used to (Tables 1 through 6).…”

“…They further speculated that α-crystallin decreases microtubule mass by increasing the critical concentration of tubulin needed for polymerization. Apparent discrepancies in how tubulin interacts with α-B crystallin may be explained by Table 4 Differentially expressed metabolic proteins in the porcine sarcoplasm in response to 12 Houck and Clark [43], who documented a nonlinear effect of α-B crystallin:tubulin molar ratios on microtubule assembly and thermal aggregation. Two cofilin 2 spots were identified, one of which (Spot 709) was increased 32 and 35% in the HS WST compared to PFTN and TN treatments, respectively.…”

Small heat shock proteins in cellular adhesion and migration

“…67 Although examples of HSPBs interacting with microtubules are scarce, a recent study showed that aB-crystallin could interact with tubulin subunits to regulate the dynamics of microtubules. 68 This interaction supposedly relies on a protein domain shared between tubulin and aBcrystallin. Such a consensus sequence is however missing in Plasmodium HSPBs.…”

Desmin and αB-crystallin interplay in the maintenance of mitochondrial homeostasis and cardiomyocyte survival

“…Previous observations suggest a physical association of desmin with mitochondria, as well as an involvement of desmin in mitochondrial homeostasis (Stromer and Bendayan, 1990;Reipert et al, 1999;Hnia et al, 2011), but the underlying mechanisms remain elusive. αΒ-crystallin associates with all three cytoskeletal networks -microfilaments (Bennardini et al, 1992;Singh et al, 2007), microtubules (Arai and Atomi, 1997;Houck and Clark, 2010) and intermediate filaments (Nicholl and Quinlan, 1994;Iwaki et al, 1989;Wisniewski and Goldman, 1998) (reviewed in Wettstein et al, 2012) -and specifically desmin (Bennardini et al, 1992;Nicholl and Quinlan, 1994;Perng et al, 1999). There is also evidence that a fraction of αΒ-crystallin is located in mitochondria (Fountoulakis et al, 2005;Martindale et al, 2005;Maloyan et al, 2005;Mitra et al, 2013); however, its role there is not completely understood.…”