Management of cytoskeleton architecture by molecular chaperones and immunophilins

Abstract

Cytoskeletal structure is continually remodeled to accommodate normal cell growth and to respond to pathophysiological cues. As a consequence, several cytoskeleton-interacting proteins become involved in a variety of cellular processes such as cell growth and division, cell movement, vesicle transportation, cellular organelle location and function, localization and distribution of membrane receptors, and cell-cell communication. Molecular chaperones and immunophilins are counted among the most important proteins that interact closely with the cytoskeleton network, in particular with microtubules and microtubule-associated factors. In several situations, heat-shock proteins and immunophilins work together as a functionally active heterocomplex, although both types of proteins also show independent actions. In circumstances where homeostasis is affected by environmental stresses or due to genetic alterations, chaperone proteins help to stabilize the system. Molecular chaperones facilitate the assembly, disassembly and/or folding/refolding of cytoskeletal proteins, so they prevent aberrant protein aggregation. Nonetheless, the roles of heat-shock proteins and immunophilins are not only limited to solve abnormal situations, but they also have an active participation during the normal differentiation process of the cell and are key factors for many structural and functional rearrangements during this course of action. Cytoskeleton modifications leading to altered localization of nuclear factors may result in loss- or gain-of-function of such factors, which affects the cell cycle and cell development. Therefore, cytoskeletal components are attractive therapeutic targets, particularly microtubules, to prevent pathological situations such as rapidly dividing tumor cells or to favor the process of cell differentiation in other cases. In this review we will address some classical and novel aspects of key regulatory functions of heat-shock proteins and immunophilins as housekeeping factors of the cytoskeletal network.

Figure 1. Model of the ATP cycle of Hsp90 and its interaction with Hop, IMMs and p23

The reversible exchange of ADP by ATP induces a conformational change in Hsp90 dimers that locks ATP into its binding site. This conformation state is stablized by the cochaperone p23, and Hsp90 recruits TPR-domain immunophilins (IMMs) and shows optimal chaperoning properties to interact with client proteins. Hsp90 returns to the ADP-bound conformation due to its intrinsic ATPase activity. Molybdate traps Hsp90 in its active conformation because the γ-phosphate is replaced by the oxyanion and prevent ADP dissociation (generating and ATP-like state). On the other hand, the benzoquinone ansamycin antibiotic geldanamycin (GA) stabilizes the ADP-bound state of Hsp90 favoring its interaction with the TPR-domain cochaperone Hop rather than with IMMs.

Figure 2. Cytoskeleton proteostasis is managed by molecular chaperones

Proteins emerging from ribosomes can be delivered to the CCT/TRiC chaperone machinery for correct folding. Those proteins that cannot be shaped properly are targeted to degradation. Based on their abundance, actin and tubulin probably occupy a substantial proportion of CCT/TRiC complexes at any given time. Native actin and tubulin assemble into microfilaments and microtubules in a nucleotide-regulated manner (ADP/ATP and GDP/GTP, respectively) with the additional assistance of other factors (not depicted here for simplicity), whereas free monomers o dimers can be targeted to proteasome degradation. Oligomeryzed small heat-shock proteins (sHsp)_n such as Hsp25/Hsp27 can be phosphorylated (asterisks) and form tetrameric structures (Phospho-sHsp)₄ able to stabilize filaments, in particular when the cells are exposed to stress. In turn, the soluble Hsp90-based heterocomplex matures in the cytosol (top of the figure) by assembling Hsp90 with Hsp70, Hsp40, p23 and a TPR-domain protein (TPR₁) that may represent the need of Hop for priming the complex. This original heterocomplex may undergo further modifications by exchanging the TPR-domain protein, for example, Hop (represented by TPR₁ in the scheme) is replaced by a high molecular weight IMM (TPR₂). The ‘mature’ heterocomplex stabilizes and also provides gain-of-function properties to several client proteins that may remain soluble or can interact with the cytoskeleton network. It should be pointed out that even the proteasome are ribosomes are also assisted by a complex of chaperones (not depicted).

Figure 3. Regulation of the Tau-microtubule interaction

(A) Schematic representation of the structural domains of the high molecular weight immunophilins FKBP51, FKBP52 and PP5 involved in Tau regulation. TPR, tetratricopeptide repeat domain or Hsp90-interacting domain; FK506, macrolide binding site to the PPIase-1 domain; HD, hinge domain (reach in polar amino acids complementary to the nuclear localization signal 1 of steroid receptors); NTP, nucleotide binding domain; CMD, calmodulin binding domain; ID, inhibitory domain. (B) Tau is phosphorylated (asterisks) in Ser and Thr residues by GSK-3b or CDK5 generating a number phospho-isoforms. Accumulation of misfolded phospho-Tau (toxic) generates aggregates as a protective response, an event that requires Hsp90 and Hsp70 and is counteracted by overexpression of FKBP52 and the Hsp90 inhibitor geldanamycin. The chaperone heterocomplex can refold Tau, whose dephosphorylated isoforms are reincorporated to microtubules. The IMM-like Ser/Thr-protein phosphatase PP5 plays a key role in this regard and is assisted by FKBP51, an IMM that enhances Tau dephosphorylation and its recycling in a PPIase-dependent manner [137]. Therefore, deficiencies in the isomerase activity of FKBP51 or in the dephosphorylation step can cause the accumulation of phosphorylated Tau protein followed by aggregation. The TPR-domain protein CHIP (C terminus of Hsc70-interacting protein) favors Tau polyubiquitination and its proteasomal degradation.

Figure 4. Subcellular redistribution of chaperones and cytoskeleton rearrangement during neuronal differentiation

(A) Images by confocal microscopy of embryonic hippocampal neurons prior and 24 h after treatment with 1 µM FK506. Image on the right hand shows a wider field. (B) Microtubules reorganize after 3 h of differentiation. (C) Intermediate filaments (IF) of FK506-differentiated neurons recruit the Hsp90-cochaperone p23.