A Tale of 12 Tails: Katanin Severing Activity Affected by Carboxy-Terminal Tail Sequences

In cells, microtubule location, length, and dynamics are regulated by a host of microtubule-associated proteins and enzymes that read where to bind and act based on the microtubule tubulin code, which is predominantly encoded in the tubulin carboxy-terminal tail (CTT). Katanin is a highly conserved AAA ATPase enzyme that binds to the tubulin CTTs to remove dimers and sever microtubules. We have previously demonstrated that short CTT peptides are able to inhibit katanin severing. Here, we examine the effects of CTT sequences on this inhibition activity. Specifically, we examine CTT sequences found in nature, alpha1A (TUBA1A), detyrosinated alpha1A, Delta 2 alpha1A, beta5 (TUBB/TUBB5), beta2a (TUBB2A), beta3 (TUBB3), and beta4b (TUBB4b). We find that these natural CTTs have distinct abilities to inhibit, most noticeably beta3 CTT cannot inhibit katanin. Two non-native CTT tail constructs are also unable to inhibit, despite having 94% sequence identity with alpha1 or beta5 sequences. Surprisingly, we demonstrate that poly-E and poly-D peptides are capable of inhibiting katanin significantly. An analysis of the hydrophobicity of the CTT constructs indicates that more hydrophobic polypeptides are less inhibitory than more polar polypeptides. These experiments not only demonstrate inhibition, but also likely interaction and targeting of katanin to these various CTTs when they are part of a polymerized microtubule filament.

Automated summary

In cells, the regulation of microtubule location, length, and dynamics is controlled by microtubule-associated proteins and enzymes that bind and act based on the microtubule tubulin code. The removal and severing of microtubules is carried out by an enzyme called katanin, which binds to the tubulin carboxy-terminal tail (CTT). Short CTT peptides have been shown to inhibit katanin severing. This study examines the effects of different CTT sequences on this inhibition activity and finds that natural CTTs have varying abilities to inhibit katanin, with beta3 CTT being ineffective. Non-native CTT tail constructs also fail to inhibit, but poly-E and poly-D peptides are able to significantly inhibit katanin.