Comparison of the density and retrograde interval velocities of WT IC-1B and the IC-1B S80 phosphomimic and dephosphomimic mutants
| Intermediate chain isoform | WT | S80A | S80D |
|---|---|---|---|
| Mean dynein density (puncta/μm of axon length) (±SEM) | 0.090 (0.007) | 0.099 (0.007) | 0.096 (0.007) |
| Mean interval retrograde velocity (μm/s) (±SEM) | 0.84 (0.02) | 0.71 (0.01) | 0.77 (0.01) |
| Number of measurements | 986 | 880 | 1328 |
| Number of puncta | 298 | 328 | 528 |
| Number of axons | 24 | 27 | 36 |
Cultured embryonic hippocampal neurons were transfected with IC-1 siRNA by electroporation, cultured on coverslips for 3 d, and then transfected with either IC-1B-mRFP wild-type, dephosphomimic S80A, or phosphomimic mutant S80D with calcium phosphate and imaged the next day. The siRNA oligonucleotide reduced the amount of endogenous IC-1 in the cells to minimize WT compensation for mutant phenotypes. There was no significant difference in the mean puncta density or interval retrograde velocity when the phospho- and dephospho-IC-1B S80 mutants were compared.