Our current model for communication along the stalk is a sliding of the helices relative to each other () by a half heptad. The proposal is that the line of hydrophobic residues in CC1 would slide in the grooves formed between the core residues in CC2. The movement would correspond to a transition between the weak binding +β registry observed in the crystal structure and the strong binding α registry trapped by SRS fusion experiments. While capable of explaining the experimental observations, there is as yet no direct proof that such a sliding occurs, nor any calculations showing that it is energetically feasible. It is perhaps useful therefore to discuss other possible mechanisms.
Fig. 4 The half-heptad sliding model of dynein stalk communication. (a) Close up view of the amino acids in the core of the stalk coiled coil (in dark blue). (b) Removal of 4 residues marked in light blue produces a construct with a 10-fold higher affinity for (more ...)
A more extreme possibility than sliding is that the coiled coil undergoes a general melting (Gee and Vallee 1998
). Such a large conformational change is reminiscent of the coiled-coil transitions occurring during the pH-dependant conformational changes observed in influenza hemagglutinin (Skehel and Wiley 2000
). The best evidence against such a model comes from electron microscopy of dynein molecules in various nucleotide states (Sale et al. 1985
; Burgess 1995
; Burgess et al. 2003
; Ueno et al. 2008
; Roberts et al. 2009
), which suggest there are no large changes in the structure of the stalk. Although the flexibility of the stalk has been observed to increase going from the apo (high affinity) to the ATP vanadate-bound (low-affinity) state (Burgess et al. 2003
), its overall dimensions remain fairly constant and consistent with that of a coiled coil (Burgess et al. 2003
; Roberts et al. 2009
). The other issue that must be addressed with a melting of the stalk is the entropic cost of unburying the hydrophobic residues in the stalk core.
It is also worth considering whether larger movements between the stalk helices should be considered in analogy to the intermolecular sliding observed between dimers of the nuclear pore protein Nup58/45 (Melcék et al. 2007
). In that case, sliding movements of ~11 Å were observed, corresponding to a whole heptad movement. However, the contacts between the α helices are entirely mediated by hydrophilic residues. The crystal structures suggested that contacts between these residues were made and broken as the helices slide past each other. These contacts appear very different from that in the dynein stalk, where the primary contribution is from hydrophobic residues. However, assuming that the half-heptad model is correct, it is porbable that the hydrophilic residues in CC1 would have to make and break contacts as CC1 and CC2 slide past each other.
Perhaps the most likely alternative to the half-heptad model is some more subtle rearrangement of the coiled coil, such as a small rotation of CC1 and CC2 with respect to each other. The best evidence for such a mechanism comes from the field of transmembrane receptors, many of which are thought to transmit signals by long-range conformational changes in the association of α helices across the plasma membrane (Matthews et al. 2006
). Hulko et al. (2006)
determined the structure of a 4-helix coiled coil called the HAMP domain that sits directly under the membrane portion of many bacterial transmembrane receptors and transmits the transmembrane signal to the cytoplasmic enzymatic domain (e.g., histidine kinase). This HAMP structure showed an unusual (knobs-to-knobs) packing that led the authors to suggest that the mechanism of signaling is the transition from this packing to the more normal knobs-in-holes variety. Such a conformational change would result in a ~28° rotation of each of the 4 helices with respect to each other. It is possible that a similar sort of rotation could occur along the length of the dynein stalk coiled coil, caused by relatively small shifts in the knobs-in-holes packing (Walshaw and Woolfson 2001
One important question is whether such relatively small changes are consistent with the SRS-fusion and crosslinking data. In the case of the SRS experiments, a large number of different registries were tested and almost all of them gave a low affinity for microtubules (with the exception of the α registry). We interpreted this to mean that if the registry was not exactly that required to produce a high affinity (α) form, then there would be a break in the registry around the fusion site and the rest of the stalk and MTBD would default to the more stable +β registry observed in the crystal structure. Such a break in a coiled coil is actually observed in an artificially shortened form of Ndc80 whose structure was recently determined (Ciferri et al. 2008
). This same explanation could clearly also be applied to smaller conformational changes, where the default conformation is favored in most constructs, whereas the high-affinity conformation is only favored in the fusions predicted to be in the α registry. In the case of the cross-linking studies (Kon et al. 2009
), it may be possible to explain the effects in terms of small conformational changes, if one assumes that the cysteines can form disulphide bonds even when they are not directly across the coiled coil from one another. Formation of diagonal cysteine crosslinks could favor one conformation of the stalk, whereas a crosslink between directly opposing cysteines would favor an alternate conformation.
In summary, in the absence of direct structural information, it is not possible to totally rule out one model of communication over another. However, one issue that is of interest is whether small conformational changes would provide a suitable mechanism for communication in dynein whose stalk, unlike the helices in transmembrane proteins, may have to bend in response to force. It may be that a half-heptad sliding mechanism provides enough of an energy barrier to prevent dynein spontaneously changing its affinity in response to force. On the other hand, given that dynein’s affinity for microtubules depends on the direction in which force is being applied (Gennerich et al. 2007
), it may be that distortion of the stalk plays a direct role in controlling dynein’s affinity for microtubules.
In all of the models for communication discussed so far, the conformational changes have occurred as a concerted movement along the whole length of the stalk. However, it is also possible that they could propagate sequentially (Carter et al. 2008
). Such a wave of rearrangements (either small rotations or larger sliding movements) may lower the overall energy barrier for the conformational change. An interesting analogy to this suggestion comes in the form of voltage sensitive ion channels, where the relative movement of the S4 helix is responsible for gating. Crystal structures of these channels show that part of S4 is in the more extended 310
helix form. This led the authors to speculate that this zone of 310
helix propagates along the S4 helix (Long et al. 2007