My long-term interest is the migration and chemotaxis
of polymorphonuclear leukocytes, the cells that provide the first
line of defense against bacterial infections. PMNs find the bacteria
through chemotaxis, directed locomotion along a chemical gradient.
(The movie attached shows PMN migration in the presence of homogeneous
chemoattractant and in the later sequences to a gradient of chemotattractant
that is first on the right side of the image and later at the
top. )Once they attach to the bacteria, they ingest and kill
them. Chemotaxis also plays an important role in development,
immune responses, wound healing and malignant metastases. Current
efforts focus on the biochemistry of the cell's motile machinery.
In particular, we are studying the signal transduction pathways
through which chemoattractants stimulate actin polymerization.
Actin
polymerization is required for cell locomotion and localized
actin polymerization is required for chemotaxis of leukocytes.
The GTPase Cdc42 induces actin polymerization
in leukocyte extracts by stimulating the Arp2/3 complex to nucleate new actin
filaments. The Cdc42-induced filaments elongate rapidly at their barbed ends.
The net result is an increase in filament number and total F-actin level. Merely
adding filaments with free barbed ends does not induce polymerization. Thus,
it appears that the Arp2/3 complex nucleated filaments are, at least transiently,
protected from capping proteins (Zigmond et al. 1998; Huang et al. 1999).
On
going studies examine various molecules that interact with
barbed ends and/or capping protein. We find a mammalian version
of CARMIL
binds capping protein
and lowers its affinity for barbed ends 10-fold. CARMIL enhances the polymerization
induced in cell extract. We find the yeast formin Bni1p nucleates new actin
filaments and binds to the filament barbed-end where it partially inhibits
elongation (Pruyne
et al. 2002 and Pring et al. 2002). Our data suggest that Bni1p is a processive
cap, moving with the barbed end as the filament elongates or shrinks. (The
animation attached shows our concept of processive capping by Bni1p).
selected
publications
Pring, M. M. Evangelista, C. Boone, C. Yang, and S.
H. Zigmond.
2003. Mechanism of formin-induced nucleation of actin filaments.
Biochemistry 42: 486-496.
Evangelista, M., S. Zigmond and C. Boone.
2003. Formins: signaling effectors for assembly and polarization
of actin filaments. J
Cell Sci 116 :2603-11
Pring M., L. Cassimeris and S. H. Zigmond.
2002 An unexplained sequestration of latrunculin A is required
in neutrophils
for inhibition of actin polymerization. Cell Motil. Cytoskel. 52:122-130.
Pruyne D., M. Evangelista, C. Yang, E. Bi, S. Zigmond,
A. Bretscher, C. Boone. (2002) Role of Formins in Actin Assembly:
Nucleation and Barbed-End Association. Science297:612-615.
Pring, M. M. Evangelista, C. Boone, C. Yang, and S. H. Zigmond 2002
Mechanism of formin-induced nucleation of actin filaments. Biochemistry avail. on line.
Zigmond, S. H. 2000. In
vitro actin polymerization using neutrophil extracts.
Regulators and Effectors of small GTPases. W. E. Balch, C.
J. Der, and A. Hall (eds.) Methods in Enzymology325:237-254.
Yang, C., M. Huang, J. DeBiasio, M.
Pring, M. Joyce, H. Miki, T.Takenawa, and S. H. Zigmond.
2000. Profilin enhances Cdc42-induced nucleation of actin polymerization. J.
Cell Biol.150:1001-1012.
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