**7. References**


ESCRT-III "spirals" are formed in the presence of these cytoskeletal elements remains a mystery. Some of these questions may be explained by recent findings that endosome delivery and fusion with the plasma membrane of the intracellular bridge may induce the disassembly of the actin cytoskeleton and lead to the secondary ingression that decreases the diameter of the intracellular bridge to ~100-200 nm (Dambournet et al., 2011; Schiel et al., 2011). Perhaps this endosome-dependent secondary ingression initiates the "de novo" recruitment of activated ESCRT-III to the abscission site by removing the actin cytoskeleton and narrowing the ICB to a smaller size (Figure 2, Endosome/ESCRT model) (Schiel and Prekeris, 2011). Indeed, while accumulation of the ESCRT-III at the abscission site can be readily detected, most recent studies have failed to observe an ESCRT "spiral" emanating from the midbody and continuing to the abscission site (Elia et al., 2011; Schiel et al., 2011).

Work from multiple laboratories in the last few years has significantly advanced our understanding about the core machinery of the abscission step of cytokinesis. All these data have demonstrated that cell abscission is an immensely complicated event that involves coordinated changes in membrane transport, microtubules, the actin cytoskeleton, septin filaments and the ESCRT complexes. How all these components are regulated, and what the mechanisms of the cross-talk between them may be, remain completely unknown and will be the focus of future studies. One of the biggest problems in studying the spatiotemporal dynamics of various cellular components during cell division has been the inability to visualize the individual organelles or cytoskeletal elements within the intracellular bridge, due to the resolution limits of the light microscopy. The emergence of novel superresolution imaging techniques, such as photo-activated localization microscopy (PALM), stimulated emission depletion microscopy (STED) and correlation high-resolution tomography will allow us to begin addressing some of these questions and

I am grateful to John Schiel (UC SOM) and Carly Willenborg (UC SOM) for a critical reading of the manuscript. I apologize to all colleagues for not being able to cite all work related to cytokinesis, due to the focused nature of this review. Work in the Prekeris laboratory has been funded by the National Institute of Health (DK064380), Susan G. Komens Breast

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Cancer Research Foundation and the Cancer League of Colorado Foundation.

**5. Conclusions and future objectives** 

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**6. Acknowledgments** 

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**9** 

*USA* 

**Enzymology and Regulation** 

*National Cancer Institute, National Institutes of Health* 

Peng Zhai, Xiaoying Jian, Ruibai Luo and Paul A. Randazzo

Arf family GTP-binding proteins, a subfamily of the Ras superfamily, are critical regulators of membrane traffic and actin remodeling (Kahn, 2009; Kahn et al., 2006; Gillingham and Munro, 2007; Donaldson and Jackson, 2011). The Arf family contains six Arf proteins in most mammals (five in humans) that are divided into three classes based on primary sequence and phylogenetic considerations (Kahn et al., 2006). The function of the Arf proteins requires switching between GDP and GTP bound forms. The accessory proteins that mediate the transitions between ArfGDP and ArfGTP function as enzymes and can be

Like other GTP binding proteins, switching between ArfGDP and ArfGTP is achieved by a controlled cycle of GTP binding and hydrolysis. The two steps are catalyzed by distinct enzymes. The conversion of ArfGDP to ArfGTP is accomplished through nucleotide exchange, with the apo form of Arf as an intermediate. Nucleotide, however, binds tightly to Arf, resulting in very slow intrinsic exchange rates, and the apo form of Arf is unstable. Guanine nucleotide exchange factors (GEFs) are enzymes that accelerate the reaction, by decreasing affinity for nucleotide and stabilizing the apo form of Arf (Casanova, 2007; Gillingham and Munro, 2007; Renault et al., 2003). Arf proteins are unusual among Ras superfamily proteins in having no detectable GTPase activity. Conversion of ArfGTP to ArfGDP is catalyzed by GTPase-activating proteins (GAPs;Gillingham and Munro, 2007; Donaldson and Jackson, 2011; Ha et al., 2008b; Kahn et al., 2008; Randazzo and Hirsch, 2004; Spang et al., 2010). The GEFs and GAPs are both large families of proteins with diverse structural features. The control of binding and hydrolysis of GTP by Arf is thought to be achieved by regulation of the ArfGAPs and ArfGEFs. The study of the ArfGAPs and ArfGEFs as allosterically controlled enzymes is providing valuable information about their

Thirty-one genes encode proteins with Arf GAP domains in humans (Kahn et al., 2008). The proteins are divided into 10 groups (Figure. 1) based on domain structure and phylogenetic analysis (Kahn et al., 2008). Six groups have the ArfGAP catalytic domain at the extreme Nterminus of the protein. In the other four groups, which comprise 20 proteins, the ArfGAP

**1. Introduction** 

studied using the formalisms of enzymology.

regulation and insights into the roles in cell physisology.

**2. ArfGAP family of proteins** 

**of ArfGAPs and ArfGEFs** 

