CLASPs Attach Microtubule Plus Ends to the Cell Cortex through a Complex with LL5β

Abstract

CLASPs are mammalian microtubule-stabilizing proteins that can mediate the interaction between distal microtubule ends and the cell cortex. Using mass spectrometry-based assays, we have identified two CLASP partners, LL5beta and ELKS. LL5beta and ELKS form a complex that colocalizes with CLASPs at the cortex of HeLa cells as well as at the leading edge of motile fibroblasts. LL5beta is required for cortical CLASP accumulation and microtubule stabilization in HeLa cells, while ELKS plays an accessory role in these processes. LL5beta is a phosphatidylinositol-3,4,5-triphosphate (PIP3) binding protein, and its recruitment to the cell cortex is influenced by PI3 kinase activity but does not require intact microtubules. Cortical clusters of LL5beta and ELKS do not overlap with focal adhesions but often form in their vicinity and can affect their size. We propose that LL5beta and ELKS can form a PIP3-regulated cortical platform to which CLASPs attach distal microtubule ends.

Talin-KANK1 interaction controls the recruitment of cortical microtubule stabilizing complexes to focal adhesions

“…The diffuse localization of the KANK1-4A-4CC mutant led to the strongly dispersed distribution of the CMSC markers ( Figure 4B,C). Furthermore, only the full-length wild type KANK1, but neither the 4A nor 4CC mutant could support efficient accumulation of CLASP2 at the peripheral cell cortex in KANK1 and KANK2-depleted cells ( Figure 4D,E), in line with the fact that cortical recruitment of CLASPs depends on LL5b (Lansbergen et al, 2006).…”

“…Due to their stereotypic round shape, HeLa cells represent a particularly convenient model for studying the impact of CMSCs on the distribution and dynamics of microtubule plus ends (Lansbergen et al, 2006;Mimori-Kiyosue et al, 2005;van der Vaart et al, 2013). In this cell line, microtubules grow rapidly in the central parts of the cell, while at the cell margin, where CMSCs cluster in the vicinity of peripheral FAs, microtubule plus ends are tethered to the cortex and display persistent but slow growth due to the combined action of several types of microtubule regulators, including CLASPs, spectraplakins and KIF21A (Drabek et al, 2006;Mimori-Kiyosue et al, 2005;van der Vaart et al, 2013).…”

“…The interaction of CLASPs with the cell cortex depends on the phosphatidylinositol 3, 4, 5-trisphosphate (PIP3)-interacting protein LL5b, to which CLASPs bind directly, and is partly regulated by PI-3 kinase activity (Lansbergen et al, 2006). Other components of the same cortical assembly are the scaffolding proteins liprin-a1 and b1, a coiled-coil adaptor ELKS/ERC1, and the kinesin-4 KIF21A (Lansbergen et al, 2006;van der Vaart et al, 2013). Both liprins and ELKS are best known for their role in organizing presynaptic secretory sites (Hida and Ohtsuka, 2010;Spangler and Hoogenraad, 2007); in agreement with this function, ELKS is required for efficient constitutive exocytosis in HeLa cells (Grigoriev et al, 2007(Grigoriev et al, , 2011.…”

“…Among them are the microtubule plus end tracking proteins (+TIPs) CLASP1/2 and the spectraplakin MACF1/ACF7, which are targeted to microtubule tips by EB1, and a homologue of EB1, EB2, which binds to mitogen-activated protein kinase kinase kinase kinase 4 (MAP4K4) (Drabek et al, 2006;Honnappa et al, 2009;Kodama et al, 2003;Mimori-Kiyosue et al, 2005). The interaction of CLASPs with the cell cortex depends on the phosphatidylinositol 3, 4, 5-trisphosphate (PIP3)-interacting protein LL5b, to which CLASPs bind directly, and is partly regulated by PI-3 kinase activity (Lansbergen et al, 2006). Other components of the same cortical assembly are the scaffolding proteins liprin-a1 and b1, a coiled-coil adaptor ELKS/ERC1, and the kinesin-4 KIF21A (Lansbergen et al, 2006;van der Vaart et al, 2013).…”

“…LL5b, liprins and ELKS form micrometer-sized cortical patch-like structures, which will be termed here cortical microtubule stabilization complexes, or CMSCs. The CMSCs are strongly enriched at the leading cell edges, where they localize in close proximity of FAs but do not overlap with them ( [Lansbergen et al, 2006;van der Vaart et al, 2013], reviewed in [Astro and de Curtis, 2015]). They represent a subclass of the previously defined plasma membrane-associated platforms (PMAPs) (Astro and de Curtis, 2015), which have overlapping components such as liprins, but may not be necessarily involved in microtubule regulation, as is the case for liprin-ELKS complexes in neurons, where they are part of cytomatrix at the active zone (Gundelfinger and Fejtova, 2012).…”