Targeted movement of cell end factors in fission yeast

Abstract

Kinesins are microtubule-based motor proteins that transport cargo to specific locations within the cell. However, the mechanisms by which cargoes are directed to specific cellular locations have remained elusive. Here, we investigated the in vivo movement of the Schizosaccharomyces pombe kinesin Tea2 to establish how it is targeted to microtubule tips and cell ends. Tea2 is loaded onto microtubules in the middle of the cell, in close proximity to the nucleus, and then travels using its intrinsic motor activity primarily at the tips of polymerizing microtubules. The microtubule-associated protein Mal3, an EB1 homologue, is required for loading and/or processivity of Tea2 and this function can be substituted by human EB1. In addition, the cell-end marker Tea1 is required to anchor Tea2 to cell ends. Movement of Tea1 and the CLIP170 homologue Tip1 to cell ends is abolished in Tea2 rigor (ATPase) mutants. We propose that microtubule-based transport from the vicinity of the nucleus to cell ends can be precisely regulated, with Mal3 required for loading/processivity, Tea2 for movement and Tea1 for cell-end anchoring.

Microtubule plus-end tracking by CLIP-170 requires EB1

“…These in vitro observations are consistent with genetic data from yeast indicating the need for both Mal3 and Tea2 for the proper cellular localization of Tip1 (29). Thus, whereas CLIP-170 is structurally similar to Tip1, the mammalian and yeast proteins differ significantly in their mechanisms of plus-end localization.…”

The EB1 Homolog Mal3 Stimulates the ATPase of the Kinesin Tea2 by Recruiting It to the Microtubule

“…Using fluorescently labeled Tea2 in live cells, it was observed that "dots" containing multipleTea2 molecules were present at the ends of MTs and that these dots moved with the growing MT ends (7). When the growing MT reached the cell end, part of the dot of Tea2 was deposited and remained attached to the cell end on retraction of the MT by depolymerization.…”

“…When the growing MT reached the cell end, part of the dot of Tea2 was deposited and remained attached to the cell end on retraction of the MT by depolymerization. Tea1, Mal3, and Tip1 also accumulate at the plus ends of growing MTs and move with the ends of polymerizing MTs to the cell tip where Tea1 and Tip1, but not Mal3, are deposited (7,12,14,15). These proteins interact physically as well.…”

“…These proteins interact physically as well. Tea2 binds to Mal3 as indicated by coimmunoprecipitation (7,8). Tea2 also binds to Tip1 through the coiled coil region of Tip1 (8), and this interaction requires the coiled coil region of Tea2.…”

“…It was first identified in a screen for morphology mutants (4) and later discovered to be identical to a kinesin gene, klp4, that was identified by a search for kinesin family members in Schizosaccharomyces pombe by reverse PCR (5). The Tea2 protein is localized in large clusters at the tips of the cell and also in punctate dots within the cell that are coincident with the ends of cytoplasmic microtubules (MTs) 1 (5,7,8). Tea2 has a conserved kinesin motor domain with an N-terminal extension of 120 amino acids and a C-terminal region that includes two predicted coiled coil regions and a C terminus that is not predicted to be coiled coil (5).…”