Slik phosphorylation of talin T152 is crucial for proper talin recruitment and maintenance of muscle attachment in Drosophila

Abstract

Talin is the major scaffold protein linking integrin receptors with the actin cytoskeleton. In Drosophila, extended Talin generates a stable link between the sarcomeric cytoskeleton and the tendon matrix at muscle attachment sites. Here, we identify phosphorylation sites on Drosophila Talin by mass spectrometry. Talin is phosphorylated in late embryogenesis when muscles differentiate, especially on T152 in the exposed loop of the F1 domain of the Talin head. Localization of a mutated version of Talin (Talin-T150/T152A) is reduced at muscle attachment sites and can only partially rescue muscle attachment compared with wild-type Talin. We also identify Slik as the kinase phosphorylating Talin at T152. Slik localizes to muscle attachment sites, and the absence of Slik reduces the localization of Talin at muscle attachment sites causing phenotypes similar to Talin-T150/ T152A. Thus, our results demonstrate that Talin phosphorylation by Slik plays an important role in fine-tuning Talin recruitment to integrin adhesion sites and maintaining muscle attachment.

The F1 loop of the talin head domain acts as a gatekeeper in integrin activation and clustering

“…Similarly the loss of kindlin-mediated control of integrin clustering by T144A/ T150A substitutions in the F1 loop (Fig. S1D) is consistent with altered muscle attachment site remodeling observed for a T152A mutant in Drosophila (Katzemich et al, 2019), and may also indicate a link between kindlin and the phosphorylation state of the F1 loop. However, independent of the regulation of the talinmembrane interaction, the identification of conserved acidic residues critical for integrin activation and clustering that are localized immediately adjacent to the inner membrane clasp reveals a much more direct way of controlling integrin activation and clustering than by differential membrane recruitment of the talin head.…”

“…Importantly, when kindlin-1 and kindlin-2 are completely removed from cells, talin no longer activates integrins and cells do not spread (Theodosiou et al, 2016). In addition, Rap1 (RAP1A and RAP1B in mammals) binding to the F0 and/or F1 subdomains of the talin head has been proposed to recruit talin to the plasma membrane, contributing to integrin activation and Rap1-mediated cell-matrix adhesion (Camp et al, 2018;Katzemich et al, 2019;Gingras et al, 2019). Thus, talin, Rap1 and kindlin play important roles in the integrin activation step, while kindlin also induces clustering of activated integrins (Feigelson et al, 2011;Schmidt et al, 2011;Feng et al, 2012;Ye et al, 2013).…”

“…Thus, it is not fully clear how the F1 loop contributes to talin-head-induced integrin activation, since it also acts downstream of the Rap1-mediated mechanism of integrin activation and membrane recruitment (Gingras et al, 2019). Nevertheless, it appears to be a site of important posttranslational modifications, because phosphorylation of the F1 loop at T144 and T150 has been associated with reduced membrane binding (Goult et al, 2010), and also affects talin recruitment to muscle attachment sites in Drosophila (Katzemich et al, 2019).…”

Genetic Control of Muscle Diversification and Homeostasis: Insights from Drosophila

“…MTJ formation is complete by the end of stage 16 and is then further refined to withstand contractile forces. Talin phosphorylation contributes to MTJ refinement [ 154 ]. This is followed by myofibril maturation and attachment to the MTJ.…”

“…Tm1 IFM isoforms are phosphorylated only in adult flies, which could have functional implications [ 260 ]. Impaired Talin phosphorylation leads to severe muscle detachment at late embryonic stages [ 154 ]. This means the right CRM regulatory mechanisms as well as post translational mechanisms such as phosphorylation [ 48 , 270 ] need to be dynamically maintained since specific protein domains are necessary for muscle specific functionality [ 269 , 271 , 272 ].…”