end3 and end4: two mutants defective in receptor-mediated and fluid-phase endocytosis in Saccharomyces cerevisiae.

Raths, Susan; Rohrer, Jack; Crausaz, Fabienne; Riezman, Howard

doi:10.1083/jcb.120.1.55

Cited by 376 publications

(356 citation statements)

References 40 publications

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“…Normally, Ste6p is then sent to the vacuole for degradation but can be trapped in the plasma membrane if a strain defective in endocytosis is analysed (Berkower et al, 1994;Kolling and Hollenberg, 1994). Reizman and co-workers have determined that the END3 gene is required for normal endocytosis to occur in Saccharomyces cerevisiae (Raths et al, 1993). To assess whether the targeting of Ycf1p to the vacuole required END3-mediated endocytosis events, Ficoll step gradients were prepared from wild-type and end3-1 cells ᮊ 1997 Blackwell Science Ltd, Molecular Microbiology, 25, 683-694 Fig.…”

Section: Vacuolar Localization Does Not Require End3-mediated Endocytmentioning

confidence: 99%

Mutational analysis of the Saccharomyces cerevisiae ATP‐binding cassette transporter protein Ycf1p

Wemmie

Moye‐Rowley

1997

Molecular Microbiology

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SummaryYcf1p is a member of the ATP-binding cassette transporter family of membrane proteins. Strong sequence similarity has been observed between Ycf1p, the cystic fibrosis transmembrane conductance regulator (CFTR) and multidrug resistance protein (MRP). In this work, we have examined the functional significance of several of the conserved amino acid residues and the genetic requirements for Ycf1p subcellular localization. Biochemical fractionation experiments have established that Ycf1p, expressed at single-copy gene levels, co-fractionates with the vacuolar membrane and that this co-fractionation is independent of vps15, vps34 or end3 gene function. Several cystic fibrosis-associated alleles of the CFTR were introduced into Ycf1p and found to elicit defects analogous to those seen in the CFTR. An amino-terminal extension shared between Ycf1p and MRP, but absent from CFTR, was found to be required for Ycf1p function, but not its subcellular localization. Mutant forms of Ycf1p were also identified that exhibited enhanced biological function relative to the wild-type protein.These studies indicate that Ycf1p will provide a simple, genetically tractable model system for the study of the trafficking and function of ATP-binding cassette transporter proteins, such as the CFTR and MRP.

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Section: Vacuolar Localization Does Not Require End3-mediated Endocytmentioning

confidence: 99%

Mutational analysis of the Saccharomyces cerevisiae ATP‐binding cassette transporter protein Ycf1p

Wemmie

Moye‐Rowley

1997

Molecular Microbiology

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“…The short actin filaments in yeast actin patches may interface with the plasma membrane through associated proteins to form a lattice that promotes membrane invagination and vesicle formation. The Arp2/3 complex nucleates actin filaments and as the filaments grow from the point of nucleation, they become attached to the plasma membrane by actin-associated endocytic accessory proteins (such as EH-domain-containing proteins and Rvs167/ amphyphysin; Raths et al, 1993;Munn et al, 1995;Santolini et al, 1999). These proteins can bind directly or indirectly to actin filaments and either to lipids in the plasma membrane or to integral membrane proteins.…”

Section: Ultrastructure Of Cortical Actin Patches In Yeastmentioning

confidence: 99%

“…myo3/5 mutant cells, which displayed numerous smaller actin patches by fluorescence microscopy ( Figure 6C, inset), did not have obvious defects in cortical actin patch ultrastructure in the unroofed spheroplasts, although the patches looked somewhat smaller in overall size ( Figure 6C). Therefore, different mutations in actinassociated proteins, which all cause defects in actin patch (Raths et al, 1993;Geli and Riezman, 1996;Warren et al, 2002), had distinct defects in actin patch organization at the resolution of electron microscopy.…”

Section: Actin Ultrastructures In Mutant Cellsmentioning

confidence: 99%

Actin and Septin Ultrastructures at the Budding Yeast Cell Cortex

Rodal

Kozubowski

Goode

et al. 2005

MBoC

174

162

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Budding yeast has been a powerful model organism for studies of the roles of actin in endocytosis and septins in cell division and in signaling. However, the depth of mechanistic understanding that can be obtained from such studies has been severely hindered by a lack of ultrastructural information about how actin and septins are organized at the cell cortex. To address this problem, we developed rapid-freeze and deep-etch techniques to image the yeast cell cortex in spheroplasted cells at high resolution. The cortical actin cytoskeleton assembles into conical or mound-like structures composed of short, cross-linked filaments. The Arp2/3 complex localizes near the apex of these structures, suggesting that actin patch assembly may be initiated from the apex. Mutants in cortical actin patch components with defined defects in endocytosis disrupted different stages of cortical actin patch assembly. Based on these results, we propose a model for actin function during endocytosis. In addition to actin structures, we found that septin-containing filaments assemble into two kinds of higher order structures at the cell cortex: rings and ordered gauzes. These images provide the first high-resolution views of septin organization in cells. INTRODUCTIONBudding yeast has been used for over 20 yr as a model organism to study cytoskeletal function because the genes encoding many components of the cytoskeleton are conserved and because yeast cells are readily amenable to parallel genetic, biochemical, and cell biological analyses. Despite rapid advances in defining the components of the yeast cytoskeleton, its ultrastructure has been difficult to image at the resolution of the electron microscope, with the exception of microtubules (O'Toole et al., 1999). This may be due to the low abundance of the cytoskeleton in yeast relative to mammalian cells, to the presence of a cell wall that complicates extraction procedures, and to high ribosome density in the yeast cytoplasm. As a result, there has been a conspicuous deficit in our understanding of the higher order organization of actin and septin polymers in vivo (Pruyne and Bretscher, 2000). Thus, although it is possible to rapidly identify and mutate cytoskeletal proteins in yeast, there is no method by which to examine the ultrastructural consequences of these mutations. This gap could be filled by developing an approach to image the yeast cytoskeleton at high resolution.The yeast actin cytoskeleton is organized into three distinct structures: motile cortical patches that are polarized in the growing bud, actin cables that run along the motherdaughter cell axis, and a contractile cytokinetic ring at the neck of dividing cells (reviewed in Pruyne and Bretscher, 2000). Cortical actin patches contain many of the actin-associated proteins common to all eukaryotes (Goode and Rodal, 2001) and are composed of actin filaments that undergo rapid turnover (Ayscough et al., 1997). Actin is essential for yeast endocytosis (Kubler and Riezman, 1993). A number of recent studies suggest that act...

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“…Ath1 could be either directly sorted into the MVB pathway at the TGN or first secreted to the plasma membrane by exocytosis and then internalized by endocytosis. To distinguish between these two possibilities, GFPAth1 was expressed in an endocytosis-deficient mutant, end3⌬, in which the internalization step of endocytosis is defective (Raths et al, 1993); Ath1 should be blocked at the plasma membrane in end3⌬ cells if it goes through the exocytic and endocytic pathways en route to the vacuole. Alternatively, if Ath1 is directly sorted into the MVB pathway, there would not be any effect on Ath1 localization in end3⌬ cells.…”

Section: Molecular Biology Of the Cell 2514mentioning

confidence: 99%

The Transmembrane Domain of Acid Trehalase Mediates Ubiquitin-independent Multivesicular Body Pathway Sorting

Huang

Reggiori

Klionsky

2007

MBoC

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Trehalose serves as a storage source of carbon and plays important roles under various stress conditions. For example, in many organisms trehalose has a critical function in preserving membrane structure and fluidity during dehydration/ rehydration. In the yeast Saccharomyces cerevisiae, trehalose accumulates in the cell when the nutrient supply is limited but is rapidly degraded when the supply of nutrients is renewed. Hydrolysis of trehalose in yeast depends on neutral trehalase and acid trehalase (Ath1). Ath1 resides and functions in the vacuole; however, it appears to catalyze the hydrolysis of extracellular trehalose. Little is known about the transport route of Ath1 to the vacuole or how it encounters its substrate. Here, through the use of various trafficking mutants we showed that this hydrolase reaches its final destination through the multivesicular body (MVB) pathway. In contrast to the vast majority of proteins sorted into this pathway, Ath1 does not require ubiquitination for proper localization. Mutagenesis analyses aimed at identifying the unknown targeting signal revealed that the transmembrane domain of Ath1 contains the information sufficient for its selective sequestration into MVB internal vesicles. INTRODUCTIONTrehalose, a nonreducing disaccharide in which two glucose molecules are connected in an ␣-1,1-glycosidic linkage, was first found in the ergot of rye in 1832 (Kopp et al., 1993). This sugar is widely distributed in various organisms, including bacteria, fungi, plants, insects, and invertebrates (Elbein, 1974). It was originally believed that trehalose served only as a sugar and energy reserve in the cell; however, it is now known that this molecule also functions as a structural component, plays a role in transport, and is involved in signaling (Elbein et al., 2003). But the most significant function of trehalose is in providing membrane protection against different kinds of stress conditions, such as heat, freezing, dehydration, anoxia, and nutrient limitation (Crowe et al., 1984).Trehalose is quite common in yeast. In Saccharomyces cerevisiae, trehalose may constitute as much as 15-20% of its dry weight when growing in a stress environment. A strong correlation has been shown between trehalose content and stress resistance (Van Dijck et al., 1995). When yeast cells grow on rich carbon sources, they have a very low level of trehalose. In contrast, as they enter the stationary phase when nutrients are exhausted or during growth on nonfermentable carbon sources, the level of this disaccharide substantially increases. Conversely, when nutrients are resupplied, trehalose is rapidly mobilized (Nwaka and Holzer, 1998). The enzymes involved in synthesis and degradation of trehalose are the key regulators of these processes.Currently the best studied pathway of trehalose biosynthesis is the one that catalyzes UDP-glucose and glucose-6-phosphate into trehalose by trehalose-6-phosphate synthase and trehalose-phosphate-phosphatase activities (Thevelein, 1984). On the other hand, the degradation...

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end3 and end4: two mutants defective in receptor-mediated and fluid-phase endocytosis in Saccharomyces cerevisiae.

Cited by 376 publications

References 40 publications

Mutational analysis of the Saccharomyces cerevisiae ATP‐binding cassette transporter protein Ycf1p

Mutational analysis of the Saccharomyces cerevisiae ATP‐binding cassette transporter protein Ycf1p

Actin and Septin Ultrastructures at the Budding Yeast Cell Cortex

The Transmembrane Domain of Acid Trehalase Mediates Ubiquitin-independent Multivesicular Body Pathway Sorting

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