We are focused on two key questions in cell biology. First,
how is the actin cytoskeleton regulated to produce mechanical
force and polarity underlying different cellular processes?
These include cell motility, endocytosis, vesicle and organelle
transport, and cytokinesis. More specifically, we are studying
how the diverse activities of numerous actin-associated
proteins are coordinated in cells to control dynamic rearrangements
of actin structures (figure 1). Second, how do the basic
cytoskeletal elements found in most eukaryotic cells (microtubules,
actin, septins, and intermediate filaments) cooperate functionally
during these same cellular processes? We are using a powerful
combination of genetic, biochemical and cell biological
approaches to address these questions in budding yeast (Saccharomyces
cerevisiae), where the core components of the cytoskeleton
are highly conserved with mammals.
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A critical breakthrough in our work was our development
of an assay to study actin assembly in yeast extracts (manuscript
in preparation). For over a decade, it has been known that
the cytoskeleton disassembles rapidly upon cell lysis in
yeast, and it has been difficult to re-stimulate actin assembly
in lysates. We now have developed a procedure to trigger
the re-assembly of > 90% of the actin into organized filamentous
structures. This method provides an unprecedented opportunity
to compare actin assembly in mutant and wild type extracts.
Further, this process can be studied in real time by spiking
reactions with rhodamine-actin and monitoring actin assembly
by fluorescence microscopy.
The actin structures reconstituted in extracts can be isolated
rapidly. This allows us to purify virtually the entire actin
cytoskeleton in a single step. Using mass spectrometry,
we have identified all of the components of this mixture,
defined the relative abundance of each actin-associated
protein in cells, and identified novel functional links
between actin and other cellular processes. Further, by
fractionating the isolated actin mixtures using gel filtration
chromatography, we have identified new activities and functional
interactions among components. For example, we showed that
the endocytic adapter protein Abp1p is a novel activator
of Arp2/3 complex, which plays a central role in promoting
actin assembly in cells (Goode et al., 2001).
Because yeast is highly amenable to genetic studies, we
can introduce mutations into any actin -associated factor
and compare its mutant activities to wild type protein.
We now are using this approach to dissect in vivo functions
and biochemical activities of each sub-unit of the Arp2/3
complex.
A long-range goal of our lab is to develop assays in extracts
to study functional interactions among the different cytoskeletal
systems. Genetic analyses have shown that cytoskeletal networks
cooperate functionally during many different cellular processes
(reviewed in Goode et al., 2000), including nuclear migration
(actin, microtubules, and intermediate filaments), organelle
inheritance (actin and intermediate filaments), and cytokinesis
(actin and septins). Our goal is to reconstitute these interactions
in cell-free extracts and identify the key factors regulating
the interactions.
Selected Publications
Chesarone M, Gould CJ, Moseley JB, Goode BL. Displacement of formins from growing barbed ends by Bud14 regulates actin network architecture and function. Dev Cell. 2009;(in press).
Chesarone MA, Goode BL. Actin nucleation and elongation factors: mechanisms and interplay. Curr Opin Cell Biol. 2009.
Stroupe ME, Xu C, Goode BL, Grigorieff N. Actin filament labels for localizing protein components in large complexes viewed by electron microscopy. RNA. 2009;15(2):244-8.
Yonetani A, Lustig RJ, Moseley JB, Takeda T, Goode BL, Chang F. Regulation and targeting of the fission yeast formin cdc12p in cytokinesis. Mol Biol Cell. 2008;19(5):2208-19. [full text in PubMed Central]
Gandhi M, Goode BL. Coronin: the double-edged sword of actin dynamics. Subcell Biochem. 2008;48:72-87.
Daugherty-Clarke K, Goode BL. WASp identity theft by a bacterial effector. Dev Cell. 2008;15(3):333-4.
Daugherty KM, Goode BL. Functional surfaces on the p35/ARPC2 subunit of Arp2/3 complex required for cell growth, actin nucleation, and endocytosis. J Biol Chem. 2008;283(24):16950-9.
Bartolini F, Moseley JB, Schmoranzer J, Cassimeris L, Goode BL, Gundersen GG. The formin mDia2 stabilizes microtubules independently of its actin nucleation activity. J Cell Biol. 2008;181(3):523-36. [full text in PubMed Central]
Sokolova O, Maiti S, Grigorieff N, Lappalainen P, Goode BL. Conformational changes in actin-binding proteins, revealed by single particle electron microscopy. Febs Journal. 2007;274:107.
Moseley JB, Bartolini F, Okada K, Wen Y, Gundersen GG, Goode BL. Regulated binding of adenomatous polyposis coli protein to actin. J Biol Chem. 2007;282(17):12661-8.
Lu J, Meng W, Poy F, Maiti S, Goode BL, Eck MJ. Structure of the FH2 domain of Daam1: implications for formin regulation of actin assembly. J Mol Biol. 2007;369(5):1258-69. [full text in PubMed Central]
Goode BL, Eck MJ. Mechanism and Function of Formins in Control of Actin Assembly. Annu Rev Biochem. 2007.
Bertling E, Quintero-Monzon O, Mattila PK, Goode BL, Lappalainen P. Mechanism and biological role of profilin-Srv2/CAP interaction. J Cell Sci. 2007;120(Pt 7):1225-34.
Okada K, Ravi H, Smith EM, Goode BL. Aip1 and Cofilin Promote Rapid Turnover of Yeast Actin Patches and Cables: A Coordinated Mechanism for Severing and Capping Filaments. Mol Biol Cell. 2006.
Moseley JB, Okada K, Balcer HI, Kovar DR, Pollard TD, Goode BL. Twinfilin is an actin-filament-severing protein and promotes rapid turnover of actin structures in vivo. J Cell Sci. 2006;119(Pt 8):1547-57.
Moseley JB, Maiti S, Goode BL. Formin proteins: purification and measurement of effects on actin assembly. Methods Enzymol. 2006;406:215-34.
Moseley JB, Goode BL. The yeast actin cytoskeleton: from cellular function to biochemical mechanism. Microbiol Mol Biol Rev. 2006;70(3):605-45.
Gandhi M, Goode BL, Chan CS. Four novel suppressors of gic1 gic2 and their roles in cytokinesis and polarized cell growth in S. cerevisiae. Genetics. 2006.
Rodal AA, Sokolova O, Robins DB, Daugherty KM, Hippenmeyer S, Riezman H, et al. Conformational changes in the Arp2/3 complex leading to actin nucleation. Nat Struct Mol Biol. 2005;12(1):26-31.
Rodal AA, Kozubowski L, Goode BL, Drubin DG, Hartwig JH. Actin and septin ultrastructures at the budding yeast cell cortex. Mol Biol Cell. 2005;16(1):372-84.
Quintero-Monzon O, Rodal AA, Strokopytov B, Almo SC, Goode BL. Structural and Functional Dissection of the Abp1 ADFH Actin-binding Domain Reveals Versatile In Vivo Adapter Functions. Mol Biol Cell. 2005;16(7):3128-39.
Moseley JB, Goode BL. Differential activities and regulation of Saccharomyces cerevisiae formin proteins Bni1 and Bnr1 by Bud6. J Biol Chem. 2005;280(30):28023-33.
D'Agostino JL, Goode BL. Dissection of Arp2/3 Complex Actin Nucleation Mechanism and Distinct Roles for Its Nucleation-Promoting Factors in Saccharomyces cerevisiae. Genetics. 2005;171(1):35-47.
View Complete Publication List on PubMed:
Bruce Goode
