Regulation of Catalytic and Non-catalytic Functions of the Drosophila Ste20 Kinase Slik by Activation Segment Phosphorylation

Panneton, Vincent; Nath, Apurba; Sader, Fadi; Delaunay, Nathalie; Pelletier, Ariane; Maier, Dominic; Oh, Karen Y.; Hipfner, David R.

doi:10.1074/jbc.m115.645952

Cited by 13 publications

(29 citation statements)

References 68 publications

(69 reference statements)

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“…The pRmHa.3puro/Slik expression construct has been described previously (Hipfner and Cohen, 2003). Sequence details of the dsRNA targeting the 5′ untranslated region (UTR) of Slik and the method for preparing it have also been described previously (Panneton et al, 2015).…”

Section: Molecular Biologymentioning

confidence: 99%

“…Anti-Slik immunoprecipitations, and immunoblotting methods have all been described previously (Panneton et al, 2015). As a substrate for Slik kinase assays, the wild-type and T152 mutant Talin FERM domains were expressed as 6xHis fusion proteins in Escherichia coli BL21 cells and purified using the B-PER 6xHis Fusion Protein Purification Kit (Thermo Fisher Scientific).…”

Section: Immunoprecipitation Mass Spectrometry and Kinase Assaysmentioning

confidence: 99%

“…As a substrate for Slik kinase assays, the wild-type and T152 mutant Talin FERM domains were expressed as 6xHis fusion proteins in Escherichia coli BL21 cells and purified using the B-PER 6xHis Fusion Protein Purification Kit (Thermo Fisher Scientific). Assays were performed as described (Panneton et al, 2015) using 1 µg of the appropriate Talin-FERM fusion protein as a substrate. Phosphorylation was detected by immunoblotting using the antiphospho-T152 Talin antiserum (generated by Genscript).…”

Section: Immunoprecipitation Mass Spectrometry and Kinase Assaysmentioning

confidence: 99%

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

et al. 2019

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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.

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Section: Molecular Biologymentioning

confidence: 99%

Section: Immunoprecipitation Mass Spectrometry and Kinase Assaysmentioning

confidence: 99%

Section: Immunoprecipitation Mass Spectrometry and Kinase Assaysmentioning

confidence: 99%

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

et al. 2019

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“…We showed that Slik was uniformly distributed at the NB cell cortex. As Slik is regulated by phosphorylation ( Panneton et al. , 2015 ), it is possible that the phosphorylated Slik is asymmetrically distributed in mitotic NBs.…”

Section: Discussionmentioning

confidence: 99%

Moesin is involved in polarity maintenance and cortical remodeling during asymmetric cell division

Abeysundara

Simmonds

Hughes

2018

MBoC

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An intact actomyosin network is essential for asymmetric cell division; however, the precise mechanisms remain unclear. We find that the actin-binding protein Moesin is required for neuroblast proliferation and mitotic progression. In addition, the asymmetric and dynamic distribution of Moesin drives the cortical remodeling of dividing neuroblasts, contributing to polarity maintenance and cell size asymmetry.

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“…The latter may also have an auto-inhibitory action because of substrate blocking and stabilisation of an inactive conformation of the αC helix, such as the case of NDR1 in its non-phosphorylated state [6]. What is more, one of the most important mechanisms related to the activation segment is auto phosphorylation, where kinases dimerize and cause phosphorylation of the activation segment [7,[8,9]. So, the exchange of activation segment has proven to be a very important action in the auto phosphorylation mechanism [11], which can occur in one or both directions [12].…”

Section: Introductionmentioning

confidence: 99%

Insights into evolutionary interaction patterns of the 'Phosphorylation Activation Segment' in kinase

et al. 2019

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We are interested in studying the phosphorylation of the kinase activation loop, distinguishing the passage from the unphosphorylated to the phosphorylated form without allostery. We performed an interaction study to trace the change of interactions between the activation segment and the kinase catalytic core, before and after phosphorylation. Results show that the structural changes are mainly due to the attraction between the phosphate group and guanidine groups of the arginine side chains of RD-pocket, which are constituted mainly of guanidine groups of the catalytic loop, the β9, and the αC helix. This attraction causes propagation of structural variation of the activation segment, principally towards the N-terminal. The structural variations are not made on all the amino acids of the activation segment; they are conditioned by the existence of two beta sheets stabilizing the loop during phosphorylation. The first,β6-β9 sheet is usually present in most of the kinases; the second, β10-β11 is formed due to the interaction between the main chain amino acids of the activation loop and the αEF/αF loop.

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Regulation of Catalytic and Non-catalytic Functions of the Drosophila Ste20 Kinase Slik by Activation Segment Phosphorylation

Cited by 13 publications

References 68 publications

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

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

Moesin is involved in polarity maintenance and cortical remodeling during asymmetric cell division

Insights into evolutionary interaction patterns of the 'Phosphorylation Activation Segment' in kinase

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