Proteomics

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U2OS Adhesion Isolations +/- CytochalasinD and Neomycin

ABSTRACT: U2OS cells preferentially utilise integrin alphaV beta5 in adhesion complexes when grown under standard cell culture conditions for 3 days. AlphaV beta5 can be found in both classical 'Focal' adhesions and in novel 'Reticular' adhesions that do not associate with F-actin or other known adhesion protein components. In order to identify components of reticular adhesions, cells were treated with cytochalasinD in order to disrupt F-actin and promote loss of focal adhesions. Remaining reticular adhesion complexes were stabilised with DTBP crosslinking reagent before adhesion complexes were isolated. Preliminary data suggested that depletion of PIP2 by Neomycin treatment would lead to disruption of reticular adhesions preferentially over focal adhesions and so this manipulation was included in these experiments.

INSTRUMENT(S): LTQ Orbitrap Elite

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Permanent Cell Line Cell, Cell Culture

DISEASE(S): Osteosarcoma

SUBMITTER: Matthew Jones  

LAB HEAD: Martin J Humphries

PROVIDER: PXD008680 | Pride | 2019-06-03

REPOSITORIES: Pride

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Publications

Reticular adhesions are a distinct class of cell-matrix adhesions that mediate attachment during mitosis.

Lock John G JG   Jones Matthew C MC   Askari Janet A JA   Gong Xiaowei X   Oddone Anna A   Olofsson Helene H   Göransson Sara S   Lakadamyali Melike M   Humphries Martin J MJ   Strömblad Staffan S  

Nature cell biology 20181022 11

Adhesion to the extracellular matrix persists during mitosis in most cell types. However, while classical adhesion complexes, such as focal adhesions, do and must disassemble to enable mitotic rounding, the mechanisms of residual mitotic cell-extracellular matrix adhesion remain undefined. Here, we identify 'reticular adhesions', a class of adhesion complex that is mediated by integrin αvβ5, formed during interphase, and preserved at cell-extracellular matrix attachment sites throughout cell div  ...[more]

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