We have sequenced three additional missense alleles of
unc-54 that suppress
unc-22 induced twitching. One,
st134, is a member of the
s74- like class while two others,
st130 and
st132 appear to define a new class of mutation in
unc-54. These latter mutations not only allow for dominant suppression of
unc-22 induced twitching, but, in addition, as homozygotes have significant A-band disorganization. The change in
st134 is TCC (ser) to TTC (phe) at position 2338, which makes this allele the 5'-most sequenced mutation in
unc-54. Most intriguing,
st134 is only two base pairs away from
s95 which is at position 2340 (i.e., these two mutations are in adjacent residues). This observation, together with the reversion studies of
s95 which we described in the last Newsletter, suggests that these two residues are essential for myosin head activity. The precise biochemical role of the region is unknown, but it is intriguing that these mutations lie very close to the putative ATP-binding domain. The change in
st132 is GAG (glu) to AAG (lys) at position 3646 and in
st130 it is TGT (cys) to TAT (tyr) at position 3739. These two mutations lie in the S1 region of the molecule. The observation that alterations in this region of the molecule can affect thick filament structure as well as the contraction/relaxation cycle is surprising. Previously, the rod has been thought to have the primary role in thick filament assembly and integrity. The stage of thick filament assembly or maintenance affected by these mutations is unknown. However, second site compensatory mutations have been isolated (see previous Newsletter) and using these, it may be possible to determine if
st130 and
st132 affect the organization of the thick filament through interactions with either the hinge or the rod portion of the molecule.