Dual specificity of SH2 domains

Credit: Vicky Summersby/NPG

Src homology 2 (SH2) domains bind to phosphotyrosine (pTyr)-containing proteins, thereby linking tyrosine kinase activity to downstream signalling cascades. Earlier studies hinted that lipids can also bind to SH2 domains. Park et al. have now characterized these interactions in detail to explain how SH2 domain-containing proteins could be regulated by plasma membrane lipids.

SH2 domains can recognize lipid headgroups specifically

Of 76 human SH2 domains analysed, 74% had high affinity for plasma membrane-mimetic (PM-mimetic) vesicles. Some of these high-affinity SH2 domains bound to phosphatidylinositol-3,4,5-trisphosphate (PtdInsP₃) and/or phosphatidylinositol-4,5-bisphosphate (PtdIns(4,5)P₂) in preference to isoelectric phosphatidylinositols, which indicates that SH2 domains can recognize lipid headgroups specifically rather than through non-specific electrostatic binding.

In addition to the pTyr-binding pocket, many SH2 domains contain alternative cationic patches (ACPs), mutation of which reduced binding to PM-mimetic vesicles and the plasma membrane localization of SH2 domains in HeLa cells. Thus, ACPs are the primary lipid-binding sites of most SH2 domains. Further studies showed that ACP morphology affects lipid binding; SH2 domains containing an ACP groove — such as the PtdInsP₃-selective T cell receptor (TCR) signalling molecule ZAP70-cSH2 — had greater phosphatidylinositol specificity than SH2 domains with a flat ACP. Moreover, ZAP70-cSH2 could bind to a lipid headgroup in the plasma membrane and a pTyr motif in the TCR ζ-chain (TCRζ) simultaneously and independently.

The authors investigated the physiological significance of this pTyr-independent, lipid-binding specificity of SH2 domains using two ZAP70-cSH2 mutants: K176E/K186E, which results in loss of PtdInsP₃ selectivity, and K206E/K251E, which reduces non-specific binding to anionic lipids, both of which have similar binding to phosphorylated TCRζ. The K206E/K251E mutant was much less effective than wild-type ZAP70-cSH2 at mediating TCR signalling throughout the whole activation period, whereas the K176E/K186E mutant was almost as active as wild-type ZAP70-cSH2 for the first few minutes but much less active during later stages. Thus, non-specific binding of ZAP70-cSH2 to membrane lipids facilitates the initial interaction with TCRζ, which is later stabilized by binding of ZAP70-cSH2 to PtdInsP₃ generated by TCR signalling. The authors propose a general model in which binding of SH2 domains to specific and non-specific lipids can modulate protein binding in a spatiotemporal manner and suggest a new strategy to control dysfunctional pTyr signalling pathways.