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Mechanobiology of Disease
Poster Abstracts
95
71-POS
Board 71
Role of Formin Fhod1 in Epec Pedestal Formation
Mrinal D. Shah
1
, G V. Shivashankar
1,4
, Alexander D. Bershadsky
1,3
, Linda J. Kenney
1,2
.
1
Mechanobiology Institute, Singapore, Singapore,
2
University of Illinois, Chicago, IL,
USA,
3
Weizmann Institute of Science, Rehovot, Israel,
4
National University of Singapore,
Singapore, Singapore.
Enteropathogenic E. coli are extracellular pathogens that cause dense puncta of polymerized
actin at the site of bacterial attachment referred to as ‘actin pedestals’. The polymerization of
actin in the pedestal is thought to be solely regulated via the Nck-WASp-Arp2/3 pathway.
Recent in vivo studies in mice have shown that Nck-deficient mice can still form actin pedestals,
suggesting a redundancy in actin polymerization pathways and the possible involvement of
multiple actin nucleators. It was therefore of interest to determine whether additional pathways
involving the formins might play a role in the formation of actin pedestals in EPEC. We
discovered that pedestal surface area was drastically reduced upon treatment with a small
molecule formin inhibitor, SMIFH2, indicating a role for formins in the pedestal formation
process. When we examined the localization of several classes of formins, we discovered a dense
localization of formins FHOD1 and mDia1 at the base of the pedestal. In agreement with our
result with formin inhibitors, we also observed that FHOD1 knockdown resulted in a similar size
reduction of pedestals and this effect was restored by complementing FHOD1 on a plasmid. We
also see interesting differences in the dynamics of Arp2/3 and FHOD1 in the pedestals, such that
very large pedestals show a near absence of Arp3 intensity but very high FHOD1 intensity. This
suggests a temporal segregation in the activity of Arp 2/3 and formins. Our findings suggest that
EPEC uses multiple pathways in the process of pedestal formation and they probably act at
different stages of pedestal the formation. We are now trying to understand the bacterial effectors
that could be involved in this differential regulation and also its importance in the pathogenesis.