ELM - Detail for LIG_IBAR_NPY

Accession:	ELME000446
Functional site class:	I-BAR domain binding site
Functional site description:	Microbial pathogens modulate the host cell actin cytoskeleton to control many processes like cell-cell contacts, movement within the cytosol, spreading to neighboring cells or development of a niche viable for intracellular bacterial replication. This is accomplished by bacterial effector proteins that hijack the major eukaryotic actin nucleators Arp2/3 complex by recruiting or mimicking nucleation promoting factors (NPFs) by a diverse array of mechanisms that serve different purposes. The major diarrhoea causing pathogen E. coli interferes with the regulation of actin signaling in host cells. A short NPY motif identified in E. coli effector protein Tir is essential for actin pedestal formation and subversion of host cell machinery.
ELM Description:	A short NPY motif present in the bacterial effector protein Tir plays an essential role in actin pedestal formation. The Tir protein is well conserved between EPEC, EHEC, and also other related bacterial pathogens. Studies have shown that the interactions between the NPY motif in Tir and the I-BAR domain of IRSp53/IRTKS is necessary for linking the bacterium to the host cell actin polymerization machinery (de Groot,2011). In the I-BAR:Tir complex (2YKT), the NPY motif interacts with a large hydrophobic-binding pocket in the central region of the I-BAR domain. The NPY motif is highly conserved and is essential for pedestal formation and none of the neighboring residues plays any critical role in pedestal formation. All three-side chains of the NPY motif interact with the I-BAR domain. The peptide backbone forms two overlapping type I β-turns (456–459, 458–461), which are stabilized by three intramolecular hydrogen bonds and an ordered water molecule. Asparagine is the most favored residue at this position since no other residue could fully satisfy the observed interactions. The pyrrolidine ring of Pro of the NPY motif rigidifies the backbone and is accommodated in a shallow hydrophobic groove on the I-BAR surface. The Tyr residue is occupied in a deeper hydrophobic pocket on I-BAR surface and substitution of this residue with Alanine virtually abolished pedestal formation by EHEC bacteria. The NPY candidate in Legionella VipA is in a predicted folded part of the sequence and has not been shown to bind with biophysical data: It is likely to be a false positive.
Pattern:	`NPY`
Pattern Probability:	`0.0000451`
Present in taxon:	Chordata
Interaction Domain:	IMD (PF08397) IRSp53/MIM homology domain (Stochiometry: 1 : 1)

I-BAR domain binding site

Microbial pathogens modulate the host cell actin cytoskeleton to control many processes like cell-cell contacts, movement within the cytosol, spreading to neighboring cells or development of a niche viable for intracellular bacterial replication. This is accomplished by bacterial effector proteins that hijack the major eukaryotic actin nucleators Arp2/3 complex by recruiting or mimicking nucleation promoting factors (NPFs) by a diverse array of mechanisms that serve different purposes. The major diarrhoea causing pathogen E. coli interferes with the regulation of actin signaling in host cells. A short NPY motif identified in E. coli effector protein Tir is essential for actin pedestal formation and subversion of host cell machinery.

ELM Description:

A short NPY motif present in the bacterial effector protein Tir plays an essential role in actin pedestal formation. The Tir protein is well conserved between EPEC, EHEC, and also other related bacterial pathogens. Studies have shown that the interactions between the NPY motif in Tir and the I-BAR domain of IRSp53/IRTKS is necessary for linking the bacterium to the host cell actin polymerization machinery (de Groot,2011). In the I-BAR:Tir complex (2YKT), the NPY motif interacts with a large hydrophobic-binding pocket in the central region of the I-BAR domain. The NPY motif is highly conserved and is essential for pedestal formation and none of the neighboring residues plays any critical role in pedestal formation. All three-side chains of the NPY motif interact with the I-BAR domain. The peptide backbone forms two overlapping type I β-turns (456–459, 458–461), which are stabilized by three intramolecular hydrogen bonds and an ordered water molecule. Asparagine is the most favored residue at this position since no other residue could fully satisfy the observed interactions. The pyrrolidine ring of Pro of the NPY motif rigidifies the backbone and is accommodated in a shallow hydrophobic groove on the I-BAR surface. The Tyr residue is occupied in a deeper hydrophobic pocket on I-BAR surface and substitution of this residue with Alanine virtually abolished pedestal formation by EHEC bacteria. The NPY candidate in Legionella VipA is in a predicted folded part of the sequence and has not been shown to bind with biophysical data: It is likely to be a false positive.

NPY

0.0000451

Chordata

IMD (PF08397) IRSp53/MIM homology domain (Stochiometry: 1 : 1)

Some pathogens have developed infection mechanisms that hijack host cell signalling so that they can extend their survival and satisfy their metabolic needs. The bacterial effector protein, Tir of Enterohemorrhagic Escherichia coli (EHEC) and Enteropathogenic E. coli (EPEC) interferes with the regulation of actin polymerization to ultimately form an actin pedestal that probably increases the contact area between the infected cell and the bacteria (Campellone,2003).

To do so, two slightly different mechanisms are used by EHEC and EPEC. Initially, the intimin receptor (Tir) is translocated by the type III secretion system (T3SS). The Tir protein contains one globular cytoplasmic N-terminal domain that is dispensable for the pedestal formation but regulates its length (Campellone,2006), one central and extracellular domain that mediates binding to the bacterial intimin protein and a C-terminal domain that differs in the different pathovars. EPEC Tir contains a short insertion around Tyr474 that is phosphorylated by host cell kinases and works as a binding motif (LIG_SH2_SRC) for the SH2 domain in Nck proteins. Nck recruited to the plasma membrane and in particular close to the bacterial attachment site, activates N-WASP by using its LIG_GBD_Chelix_1 motif to bind to the GBD domain of N-WASP. The VCA regions can then activate Arp2/3 and initiate actin polymerization leading to pedestal formation.

EHEC Tir lacks the Nck binding property; instead, it sequesters and activates N-WASP by a ternary complex that includes a second effector protein, EspFU, and the host proteins IRSp53 or IRTKS. EHEC Tir uses the NPY motif located at the C-terminal domain to bind to the I-BAR domain of IRSp53/IRTKS (Campellone,2006; Brady,2007; Weiss,2009). IRSp53/IRTKS has an SH3 domain that interacts with the multiple LIG_SH3_3 motifs in EspFU. EspFU, in addition, has a LIG_GBD_Chelix_1 that mimics the same motif in Nck then converging in the activation of actin polymerization and pedestal formation with EPEC.

Another disease-causing bacteria, Legionella pneumophila, uses the effector protein VipA as a link between actin dynamics and the endocytic pathway, with its COOH-terminal region mediating the interaction with the cytoskeleton component and the NH2 region with the endocytic vesicles. The NPY motif within the NH2 region was reported to be necessary for interference with organelle trafficking in a yeast infection model (Bugalhao,2016); it remains to be proven that the motif in VipA mediates the binding to the I-BAR domain because it is in a region predicted to be folded. At time of entry preparation, no experimentally verified host cellular proteins are identified: Sequence-based prediction shows some plausible proteins like shank2, myosin15 and Clasp1. The co-occurrence of shank family proteins and IRSp53 at the post synaptic density (Dosemeci,2017) might support the prediction.

Biological Process:
Pathogenesis (also annotated in these classes: )
Cellular Compartment:
Cytoplasmic Side Of Membrane (also annotated in these classes: )
Cytosol (also annotated in these classes: )
Molecular Function:
Protein Binding (also annotated in these classes: )

Acc., Gene-, Name	Start	End	Subsequence	Logic	#Ev.	Organism	Notes
O60241 ADGRB2 AGRB2_HUMAN	1417	1419	GPAYGSLQNPYGMTFQPPPP	TP	1	Homo sapiens (Human)
Q9UPX8 SHANK2 SHAN2_HUMAN	608	610	NKRGQMPENPYSEVGKIASK	TP	1	Homo sapiens (Human)
Q7Z460 CLASP1 CLAP1_HUMAN	1265	1267	PGPRARDYNPYPYSDAINTY	TP	1	Homo sapiens (Human)
Q9UKN7 MYO15A MYO15_HUMAN	423	425	PPPIPSPHNPYAHAMDDIAE	TP	1	Homo sapiens (Human)
Q5C8M7 vipA Q5C8M7_LEGPN	76	78	TVLLSAKTNPYLLDCWISFI	FP	1	Legionella pneumophila
C6UYL8 tir TIR_ECO5T	456	458	TSSIGTVQNPYADVKTSLHD	TP	13	Escherichia coli O157:H7 str. TW14359	2YKT 2YKT 2
B7UM99 tir TIR_ECO27	452	454	SDSSSEVVNPYAEVGGARNS	TP	2	Escherichia coli O127:H6 str. E2348/69

Acc., Gene-, Name

Start

End

Subsequence

Logic

#Ev.

Organism

Notes

O60241 ADGRB2
AGRB2_HUMAN

1417

1419

GPAYGSLQNPYGMTFQPPPP