THE JOURNAL OF BIOLOGICAL CHEMISTRY

Vol. 276, No. 23, Issue of June 8, pp. 19679 19682, 2001
Printed in U.S.A.

Minireview
The Yeast Cell Wall and Septum
as Paradigms of Cell Growth
and Morphogenesis*
Published, JBC Papers in Press, April 17, 2001,
DOI 10.1074/jbc.R000031200
Enrico Cabib, Dong-Hyun Roh, Martin Schmidt,
Luciana B. Crotti, and Archana Varma
From the Laboratory of Biochemistry and Genetics,
NIDDK, National Institutes of Health,
Bethesda, Maryland 20892

Composition and Structure of the Yeast Cell Wall

The yeast cell wall consists mainly of polysaccharides made
up of three sugars, glucose, mannose, and N-acetylglucosamine. One of the glucose polysaccharides, (133)glucan, is
the major structural component of the wall. Another,
(136)glucan, is relatively minor in amount but very important for cross-linking (see below). The mannose polysaccharides are linked to proteins to form a mannoprotein layer mainly
localized at the external surface and acting as a filter for large
molecular weight materials (2). Finally, the small amount of
GlcNAc present in the wall (13%) is in the (134)-linked
linear polysaccharide chitin, which is mostly concentrated in
the septal region although some is dispersed throughout the
wall. Despite its small quantity, chitin is essential for yeast
survival (3), probably because of its central role in septation.
An early indication that the yeast cell wall components are
* This minireview will be reprinted in the 2001 Minireview Compendium, which will be available in December, 2001.
To whom correspondence should be addressed: NIDDK, National
Institutes of Health, Bldg. 8, Rm. 403, Bethesda, MD 20892. Tel.:
301-496-1008; Fax: 301-496-9431; E-mail: enricoc@bdg10.niddk.nih.
gov.
This paper is available on line at http://www.jbc.org

Biosynthesis of (133)Glucan as a Model for

Cell Wall Synthesis and Its Regulation
The growth of the cell wall must accompany that of the cell
that it encloses. In particular, new cell wall must be generated
at the emergence of a new bud; its synthesis continues throughout the growth of the bud and stops at maturation. To study the
temporal and spatial regulatory controls that direct this process, (133)glucan, the major structural component of the yeast
cell wall, has been used as a model. The enzymatic system that
catalyzes the synthesis of (133)glucan from UDP-glucose is
bound to the plasma membrane (for review, see Ref. 11). An
early finding was that GTP is a potent activator of (133)glucan synthase at micromolar concentration (11). Later studies
with yeast and other fungi led to the stepwise solubilization of
two fractions from the membrane, both required simultaneously for glucan synthesis. One contains a small (26 kDa)

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Fungal cells, like their bacterial and plant counterparts but

unlike animal cells, are encased in a sturdy shell, the cell wall,
which is essential for their survival. Fungal cells have a very
high turgor pressure, and even a minor chink in the cell wall
structure can lead to bursting and death (1). Yet, because the
cell wall is external to the cell, it must grow with it and endure
all the changes that the cell undergoes during the division
cycle. To ensure the continuous integrity of the cell wall without interfering with its plasticity and constant growth, elaborate control mechanisms must be operating, which need to be
strictly coordinated with those governing the cell cycle.
There are two major reasons for studying the fungal cell wall
and its specialized structure for cell division, the septum. 1) Of
theoretical interest is that the cell wall, because it imparts
shape to the cell in a constantly changing pattern and because
of its relatively simple composition, serves as a good model for
morphogenesis at the molecular level. 2) Another reason, of an
applied nature, is that cell wall synthesis is an attractive target
for antifungal agents because it is unique to the pathogen.
Because of the versatility of budding yeast for biochemical
and genetic studies, the most comprehensive body of work on
fungal cell walls has been carried out with this organism.
Therefore, this review will deal specifically with Saccharomyces cerevisiae.

attached to each other by covalent linkages was that mannoproteins, insoluble in native walls, became soluble in water
after digestion of the walls with an endo-(133)-glucanase (2).
This suggested a direct or indirect attachment between
(133)glucan and mannoproteins. The first fully characterized
linkage between cell wall components was the (134) bond
between the reducing terminal GlcNAc of a chitin chain and
the nonreducing terminal glucose of (133)glucan (Fig. 1A) (4).
Other studies revealed an association between mannoproteins,
(136)glucan and (133)glucan (5). Finally, the linkages of a
complex including those three components plus chitin were
identified and partially characterized (Fig. 1B) (6). In this
complex, (136)glucan occupies a central position. To it, chitin
and (133)glucan are directly attached by glycosidic linkages,
whereas mannoprotein is joined to the polysaccharide through
the lipidless remnant of a glycosylphosphatidylinositol anchor
(Fig. 1B). These anchors have been identified as signals for the
incorporation of proteins in the cell wall after transit through
the plasma membrane (for review, see Ref. 7).
In addition to the complex shown in Fig. 1B, another was
isolated that lacked the mannoprotein. Together with the previous identification of a chitin-(133)glucan compound, this
finding suggested the notion of a flexible building block (6) or
module (7) for cell wall construction. These modules, of somewhat variable structure, would form a lattice all around the
cell, thus generating an essentially continuous envelope. The
modules may be joined to each other through hydrogen bonds
(Fig. 1C). It is also possible that the (133)glucan branches
joined to the main chain by (136) linkages are longer than
formerly thought, and this would provide the means for a
practically endless network of the branched polysaccharide
around the cell (Fig. 1D).
Other linkages are also present in the cell wall. A family of
proteins called Pir (proteins with internal repeats) seems to be
attached directly to (133)glucan through an alkali-labile
linkage of as yet unresolved nature (8, 9). In addition, some
proteins are released from the wall with sulfhydryl compounds
(Fig. 1, C and D) (10).
Obviously, the cross-links must contribute to the mechanical
strength of the wall. They could also be important in the loosening and remodeling of the wall, such as at bud emergence.
Some critical linkages could be cut temporarily by appropriate
enzymes to allow for wall expansion, while the remainder of the
network would keep the wall together.

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Minireview: Yeast Cell Wall and Septum as Paradigms

GTP-binding protein; the other harbors the catalytic component(s) of the synthase (11). The GTP-binding protein was
identified as Rho1p (12, 13), whereas a large hydrophobic protein, Fks1p, was found in the catalytic fraction (14). Properties
of mutants and experiments with specific inhibitors indicate
that Fks1p or its close homolog Fks2p is necessary for
(133)glucan synthesis in vivo (14). There is no proof that
Fks1p or Fks2p contains the catalytic site for glucan synthesis,
but they are the only present candidates for such a function.
Their participation in the synthetic reaction is supported by the
coimmunoprecipitation of Fks1p and Rho1p (13).
The discovery that a GTP-binding protein was the regulator
of (133)glucan synthase went a long way to explain how the
synthesis of the polysaccharide product can be started or shut
off (11). GTP-binding proteins can act as molecular switches
because they are active when bound to GTP and inactive when
bound to GDP. The conversion from active to inactive form is
stimulated by GTPase-activating proteins, whereas the opposite shift is helped by GTP-GDP exchange factors. Two proteins, Rom1p and Rom2p, have been identified as GTP-GDP
exchange factors for Rho1p (15), whereas Bem2p and Sac7p are
potential, albeit yet unproven, GTPase-activating proteins for
Rho1p (11).
The shift of Rho1p between the active and inactive state
provides a rationale for the switching on and off of (133)glucan and cell wall synthesis. As pointed out below, Rho1p has
other functions in addition to regulation of (133)glucan synthase. We recently found that rho1 mutants with a temperature-sensitive defect specific for the glucan synthesis function
die at the nonpermissive temperature.1 Although for some time
rho1 mutants incorporate other components in the cell wall,
these components cannot be anchored to (133)glucan. These
results underline the importance of (133)glucan for the preservation of cell wall architecture.

Biosynthesis of Other Cell Wall Components

The mechanism of biosynthesis of (136)glucan, a central
component of the wall cross-linked modules, has been studied
by a genetic approach. Bussey and co-workers (reviewed in Ref.
1

D.-H. Roh and E. Cabib, unpublished results.

Localization of Cell Wall Growth

In addition to temporal switches of the type involving Rho1p
and (133)glucan synthase, spatial controls are required to
confine growth of the cell wall to the bud and to direct it in such
a way that a bud will be made with the correct shape. These
controls are part of the processes of cell polarization and actin
cytoskeleton organization, which are beyond the scope of this
article (for reviews see Refs. 22 and 23). Suffice it to say that
immediately after emergence, the bud grows at the tip (apical
growth). Accordingly, Rho1p (13, 24) and Fks1p (13), components of the (133)glucan complex, localize at that site (Fig.
2A). Growth then becomes more uniformly distributed (isotropic growth, Fig. 2B) to generate the ellipsoidal shape of the
daughter cell. These changes in growth pattern are accompanied by concomitant changes in the distribution of actin
patches (23). Another component of the actin cytoskeleton that
is essential for growth, actin filaments, appears to direct into
the bud the secretory vesicles that carry cell wall and membrane components as well as enzymes for cell wall synthesis
(23).

The Preservation of Cell Wall Integrity

As mentioned above, it is imperative for the cell to maintain
cell wall integrity at all times, including stages of the cell cycle
involving extensive remodeling of the wall, such as budding
and cell division, and situations of stress that may arise in
nature, such as extremes of temperature and pH. The cell
receives signals of the stressful conditions through a battery of
sensors (the proteins Wsc1p/HCS77, Wsc2p, Wsc3p, and
Mid2p, at present count) bound to the plasma membrane (25
28). The signal is transmitted somehow to Rho1p (25), which, in
addition to its function in (133)glucan synthesis, acts as an
activator of the protein kinase C, Pkc1p (29, 30). Pkc1p, in turn,
controls a mitogen-activated protein kinase cascade, which is
required for cell wall integrity (31). Defects in Pkc1p and in
some members of the cascade lead to cell lysis. At least one of
the functions of the mitogen-activated protein kinase cascade is
as a positive regulator of the transcription of several genes
engaged in cell wall biosynthesis, including FKS1 (32). Thus,
Rho1 acts on (133)glucan and cell wall formation both directly (as a component of (133)glucan synthase) and indirectly (as an activator of Pkc1p).

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FIG. 1. Architecture of the yeast cell wall. A, the linkage between

chitin and (133)glucan. B, structure of a complex that contains all
four components of the cell wall, (133)glucan, (136)glucan, mannoprotein, and chitin. Arrows point to reducing ends. P is a phosphate
group. C and D, two possible ways of attaching together cell wall
modules, either by hydrogen bonds (sets of three black bars in C) or by
(133)glucan side chains (D). The color code is the same as in B.

16) took advantage of the fact that (136)glucan is a receptor

for the K1 killer toxin to isolate mutants resistant to the toxin
because of their diminished (136)glucan content. Several
genes required for synthesis of the polysaccharide were cloned
by complementation of the mutations. Based on the sequences
of the corresponding proteins, guesses were made about their
intracellular localization. From this evidence it was concluded
that the synthesis of (136)glucan probably begins in the
endoplasmic reticulum, continues in the Golgi, and is completed at the cell surface (16). However, Montijn et al. (17) were
unable to find intracellular material reacting with a
(136)glucan antibody, even in mutants that accumulated
secretory vesicles. Thus, the mechanism of synthesis of
(136)glucan remains obscure.
As for mannoproteins, a large amount of information has
been built up over the years about the biochemical steps
through which oligosaccharides are constructed and transferred to protein in the ER, followed by further elongation in
the Golgi (see Refs. 18 21 for reviews). On the other hand,
little is known about the mechanism and sites of incorporation
of mannoproteins in the cell wall, including the putative transglycosylation reactions by which the glycosylphosphatidylinositol anchors are attached to (136)glucan with elimination of
their lipid moiety.

Minireview: Yeast Cell Wall and Septum as Paradigms

19681

FIG. 2. Cell wall growth and septum formation. The cell wall is in
gray, and the plasma membrane is the red broken line. In A, CSIII
(black), at the base of an emerging bud, has catalyzed the synthesis of
the chitin ring (green). Bud growth is in the apical phase (arrow), and
Rho1p and Fks1p, components of (133)glucan synthase, are at the
bud tip. B, as the bud enlarges, the growth becomes isotropic (arrows).
Rho1p and Fks1p are no longer detectable. Notice in A and B the chitin
(green dashes) interspersed in the cell wall of the mother cell but not in
the bud. C, a contractile ring (CR) invaginates the plasma membrane at
cytokinesis. Concomitantly, CSII (brown) catalyzes the synthesis of
primary septum chitin (green). D, the primary septum is now complete
and secondary septa (SS, yellow) have been laid down from the mother
and daughter cell. Chitin appears in the daughter cell wall. E, cell
separation is facilitated by a partial digestion of the primary septum by
a chitinase (Chit). CSI (purple) serves as a repair enzyme to provide
extra chitin if chitinase action goes too far. BS, bud scar.

The Septum, a Specialized Structure

of the Cell Wall
At cell division, a cross-wall, the septum, is made between
the mother and daughter cell to permit their separation without lysis (reviewed in Refs. 33 and 34). The yeast septum is
made in three different stages. The first takes place very early
when a chitin ring is formed at the basis of an emerging bud
(Fig. 2A). The second occurs at cytokinesis, when a contractile
actomyosin ring leads to the invagination of the plasma membrane (35, 36). As the membrane invaginates, chitin is secreted
from it to form a disc, the primary septum, that is the first
structure separating the dividing cells (Fig. 2C). Once the primary septum is completed, the third and last stage starts with
the buildup by mother and daughter cells of secondary septa
that appear to have a similar composition to the remainder of
the cell wall (Fig. 2D). Finally, the two cells are separated by
the action of a chitinase (37) that partially hydrolyzes the
primary septum (Fig. 2E).
Because of its simple geometrical shape and of its importance
for cell division, the septum is an attractive structure for morphogenetic studies. Most such studies have dealt with the
biosynthesis of chitin because of the central role of this polysaccharide in septum formation. On the other hand, little is known
about the formation of secondary septa, and they will not be
discussed here. Chitin synthesis is catalyzed by synthases that
transfer GlcNAc from UDP-GlcNAc to a growing polysaccha-

Function of Chitin Synthases and Their Regulation

Examination of the phenotype of mutants in the three chitin
synthase genes led to an understanding of their function (3).
chs2 mutants are unable to construct a primary septum. Although they usually succeed in completing cell division, their
septa are thick, amorphous, and often show lacunae. Chitinase
cannot act on these septa, and as a result cell separation is
defective and the cells form large clumps. It was concluded that
CSII is responsible for the synthesis of primary septum chitin.
chs3 mutants do not lay down the chitin ring at the base of
emerging buds, but they have a tri-layered septum. They also
lack the chitin dispersed in the cell wall and involved in crosslinks (4). As for CSI, the defect in this enzyme had a more
subtle phenotype; when grown in a medium with poor buffering
power, many daughter cells lyse during cell separation, with a
small hole in the center of the birth scar (1). Chitinase inhibitors reduced the defect and deletion of the chitinase gene abolished it (45). Therefore it appears that CSI has a repair function by providing replacement chitin when the chitinase
involved in cell separation (see above) is activated by the decreasing pH of the medium and digests too much of the primary
septum.
It is interesting that although single mutants in each one of
the three chitin synthase genes, CHS1, CHS2, and CHS3, as
well as double mutants chs1 chs2 and chs1 chs3 are viable,
simultaneous mutation of CHS2 and CHS3 is lethal (3). The
reason for this synthetic lethality is not well understood, but
the result indicates that chitin is essential for cell survival and
therefore a potential target for antifungals.
How do the three chitin synthases carry out their different
tasks? We will start with CSIII because more is known about
this synthase. The reason for this resides in the discovery that
Calcofluor White, a fluorescent dye that binds to chitin, induces
a great increase in the synthesis of the polysaccharide and
arrests cell growth. Mutations in CHS3 or other genes required
for Chs3p function will result in resistance to Calcofluor. With
2
The abbreviations used are: CSI, CSII, CSIII, chitin synthase I, II,
and III, respectively; ER, endoplasmic reticulum. Roman numerals are
used to designate the enzymes because, in addition to the putative
catalytic moieties, i.e. Chs1p, Chs2p and Chs3p, they may contain
activators (such as Chs4p for CSIII) and perhaps proteins needed to
guide the newly formed polysaccharide through the plasma membrane.

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ride chain. Three chitin synthases, CSI,2 CSII, and CSIII, have
been identified in yeast, and the genes believed to code for their
catalytic components, CHS1, CHS2, and CHS3, have been
cloned (33, 34). The corresponding proteins show a high level of
identity in the middle and carboxyl-terminal portion, whereas
a nonessential amino-terminal region of variable length is
quite dissimilar (38). Several mutations of Chs2p that lead to
inactive proteins have been identified (39, 40). The shared
motif QRRRW seems to be essential for activity and function of
all three synthases (41).
All three chitin synthases are membrane-bound, in part to
the plasma membrane and in part to vesicles of uncertain
origin that have been named chitosomes (33, 42). A general
feature of fungal chitin synthases, shared by CSI and CSII, is
that they are found in a mostly inactive state. Partial proteolysis, usually by trypsin, leads to a striking increase in activity
(33). The latent form of the synthases has been termed the
zymogen. CSIII is isolated in an active state, apparently because an activator, most probably Chs4p, is bound to it (43, 44).
Extraction with detergents or mutations in CHS4 revealed a
zymogenic form also for CSIII (43). How CSI and CSII are
activated in vivo is unknown, but the very existence of the
zymogenic forms suggests that activation of the zymogen must
be an important step in their regulation.

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Minireview: Yeast Cell Wall and Septum as Paradigms

Concluding Remarks
The cell wall and septum together make up about one-third
of the yeast cell dry weight. Thus, the cell devotes a great deal
of its energy and of its synthetic apparatus to generate these
structures. Moreover, cell wall and septum construction is part
of cell growth and division; therefore, it must be strictly regulated in space and time to accompany the other processes that
are part of the cell cycle. By studying those regulatory mechanisms we can learn how the cell localizes proteins, activates
enzymes, and builds structures with a defined shape. Thus, the
study of yeast cell wall and septum formation has provided a
rich harvest of knowledge about fundamental cellular processes. At the same time, new targets for antifungal compounds
have been defined. The main ones so far have been chitin and
(133)glucan synthesis, for which inhibitors, such as the
polyoxins and nikkomycins for chitin and the semisynthetic
echinocandins for glucan, have become available. The glucan
synthesis inhibitors are now undergoing clinical tests. Further
study of the fungal cell wall may be expected to continue to
furnish both exciting new findings and clinical applications.
AcknowledgmentWe thank Jan Wolff for a critical reading of the
manuscript.
REFERENCES
1. Cabib, E., Sburlati, A., Bowers, B., and Silverman, S. J.(1989) J. Cell Biol. 108,
16651672

2. Zlotnik, H., Fernandez, M. P., Bowers, B., and Cabib, E.(1984) J. Bacteriol.
159, 1018 1026
3. Shaw, J. A., Mol, P. C., Bowers, B., Silverman, S. J., Valdivieso, M. H., Duran,
A., and Cabib, E. (1991) J. Cell Biol. 114, 111123
4. Kollar, R., Petrakova, E., Ashwell, G., Robbins, P. W., and Cabib, E.(1995)
J. Biol. Chem. 270, 1170 1178
5. Kapteyn, J. C., Montijn, R. C., Vink, E., de la Cruz, J., Llobell, A., Shimoi, H.,
Lipke, P., and Klis, F. M.(1996) Glycobiology 6, 337345
6. Kollar, R., Reinhold, B. B., Petrakova, E., Yeh, H. J. C., Ashwell, G., Drgonova,
J., Kapteyn, J. C., Klis, F. M., and Cabib, E.(1997) J. Biol. Chem. 272,
1776217775
7. Lipke, P. N., and Ovalle, R.(1998) J. Bacteriol. 180, 37353740
8. Mrsa, V., Seidl, T., Gentzsch, M., and Tanner, W.(1997) Yeast 13, 11451154
9. Kapteyn, J. C., Van Egmont, P., Slevi, E., Van Den Ende, H., Makarow, M.,
and Klis, F. M.(1999) Mol. Microbiol. 31, 18351844
10. Moukadiri, I., Jaafar, L., and Zueco, J.(1999) J. Bacteriol. 181, 4741 4745
11. Cabib, E., Drgonova, J., and Drgon, T.(1998) Annu. Rev. Biochem. 67, 307333
12. Drgonova, J., Drgon, T., Tanaka, K., Kollar, R., Chen, G.-C., Ford, R. A., Chan,
C. S. M., Takai, Y., and Cabib, E.(1996) Science 272, 277279
13. Qadota, H., Python, C. P., Inoue, S. B., Arisawa, M., Anraku, Y., Zheng, Y.,
Watanabe, T., Levin, D. E., and Ohya, Y.(1996) Science 272, 279 281
14. Douglas, C. M., Foor, F., Marrinan, J. A., Morin, N., Nielsen, J. B., Dahl, A. M.,
Mazur, P., Baginski, N., Li, W., El-Sherbeini, M., Klemas, J. A., Mandala,
S. M., Frommer, B. M., and Kurtz, M. B., (1994) Proc. Natl. Acad. Sci.
U. S. A. 91, 1290712911
15. Ozaki, K., Tanaka, K., Imamura, H., Hihara, T., Kameyama, T., Nonaka, H.,
Hirano, H., Matsuura, Y., and Takai, Y.(1996) EMBO J. 15, 2196 2207
16. Shahinian, S., and Bussey, H. (2000) Mol. Microbiol. 35, 477 489
17. Montijn, R. C., Vink, E., Muller, W. H., Verkleij, A. J., Van Den Ende, H.,
Henrissat, B., and Klis, F. M.(1999) J. Bacteriol. 181, 7414 7420
18. Burda, P., and Aebi, M.(1999) Biochim. Biophys. Acta 1426, 239 257
19. Knauer, R., and Lehle, L.(1999) Biochim. Biophys. Acta 1426, 259 273
20. Dean, N.(1999) Biochim. Biophys. Acta 1426, 309 322
21. Strahl-Bolsinger, S., Gentzsch, M., and Tanner, W.(1999) Biochim. Biophys.
Acta 1426, 297307
22. Pruyne, D., and Bretscher, A. (2000) J. Cell Sci. 113, 365375
23. Pruyne, D., and Bretscher, A. (2000) J. Cell Sci. 113, 571585
24. Yamochi, W., Tanaka, K., Nonaka, H., Maeda, A., Musha, T., and Takai,
Y.(1994) J. Cell Biol. 125, 10771093
25. Verna, J., Lodder, A., Lee, K. M., Vagts, A., and Ballester, R.(1997) Proc. Natl.
Acad. Sci. U. S. A. 94, 13804 13809
26. Gray, J. V., Ogas, J. P., Kamada, Y., Stone, M., Levin, D. E., and Herskowitz,
I.(1997) EMBO J. 16, 4924 4937
27. Rajavel, M., Philip, B., Buehrer, B. M., Errede, B., and Levin, D. E.(1999) Mol.
Cell. Biol. 19, 3969 3976
28. Ketela, T., Green, R., and Bussey, H.(1999) J. Bacteriol. 181, 3330 3340
29. Nonaka, H., Tanaka, K., Hirano, H., Fujiwara, T., Kohno, H., Umikawa, M.,
Mino, A., and Takai, Y.(1995) EMBO J. 14, 59315938
30. Kamada, Y., Qadota, H., Python, C. P., Anraku, Y., Ohya, Y., and Levin,
D. E.(1996) J. Biol. Chem. 271, 91939196
31. Gustin, M. C., Albertyn, J., Alexander, M., and Davenport, K.(1998) Microbiol.
Mol. Biol. Rev. 62, 1264 1300
32. Igual, J. C., Johnson, A. L., and Johnston, L. H.(1996) EMBO J. 15, 50015013
33. Cabib, E., Shaw, J. A., Mol, P. C., Bowers, B., and Choi, W.-J.(1996) in The
Mycota (Bramble, R., and Marzluf, G. A., eds) Vol. III, pp. 243267, Springer-Verlag, Berlin
34. Cid, V. J., Duran, A., del Rey, F., Snyder, M. P., Nombela, C., and Sanchez,
M.(1995) Microbiol. Rev. 59, 345386
35. Epp, J. A., and Chant, J.(1997) Curr. Biol. 7, 921929
36. Lippincott, J., and Li, R.(1998) J. Cell Biol. 140, 355366
37. Kuranda, M. J., and Robbins, P. W.(1991) J. Biol. Chem. 266, 19758 19767
38. Ford, R. A., Shaw, J. A., and Cabib, E.(1996) Mol. Gen. Genet. 252, 420 428
39. Nagahashi, S., Sudoh, M., Ono, N., Sawada, R., Yamaguchi, E., Uchida, Y.,
Mio, T., Takagi, M., Arisawa, M., and Yamada-Okabe, H.(1995) J. Biol.
Chem. 270, 1396113967
40. Yabe, T., Yamada-Okabe, T., Nakajima, T., Sudoh, M., Arisawa, M., and
Yamada-Okabe, H.(1998) Eur. J. Biochem. 258, 941947
41. Cos, T., Ford, R. A., Trilla, J. A., Duran, A., Cabib, E., and Roncero, C.(1998)
Eur. J. Biochem. 256, 419 426
42. Chuang, J. S., and Schekman, R. W.(1996) J. Cell Biol. 135, 597 610
43. Choi, W.-J., Sburlati, A., and Cabib, E.(1994) Proc. Natl. Acad. Sci. U. S. A. 91,
4727 4730
44. Trilla, J. A., Cos, T., Duran, A., and Roncero, C.(1997) Yeast 13, 795 807
45. Cabib, E., Silverman, S. J., and Shaw, J. A.(1992) J. Gen. Microbiol. 138,
97102
46. Trilla, J. A., Duran, A., and Roncero, C.(1999) J. Cell Biol. 145, 11531163
47. Santos, B., and Snyder, M.(1997) J. Cell Biol. 136, 95110
48. Ziman, M., Chuang, J. S., Tsung, M., Hamamoto, S., and Schekman, R.(1998)
Mol. Biol. Cell 9, 15651576
49. De Marini, D. J., Adams, A. E. M., Fares, H., De Virgilio, C., Valle, G., Chuang,
J. S., and Pringle, J. R.(1997) J. Cell Biol. 139, 7593
50. Ziman, M., Chuang, J. S., and Schekman, R. W. (1996) Mol. Biol. Cell 7,
1909 1919
51. Choi, W.-J., Santos, B., Duran, A., and Cabib, E.(1994) Mol. Cell. Biol. 14,
76857694

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this screen and with another screen based on glucosamine

incorporation, CHS3 itself and four other genes, now called
CHS4 to CHS7, were cloned (see Ref. 34 for review). One
question is how Chs3p reaches the site(s) where it exerts its
function. This journey starts in the ER. For export of Chs3p
from the ER, another protein, Chs7p, is required (46). The next
station is the Golgi, where Chs3p appears to be associated with
a different protein, Chs5p (47). Chs5p and Chs6p (48) are
necessary for the next step, which is transfer to vesicles (chitosomes) that will carry Chs3p to the plasma membrane (42,
48). However, Chs3p is not accompanied by Chs5p to its final
destination, a ringlike area at the base of an emerging bud,
where the chitin ring is laid down (42, 47) (Fig. 2A). Chs3p may
be retained in this location through its interaction with the
putative activator, Chs4p. The latter can bind to Bni4p, which
in turn can attach to the septin ring (49), a microfilament ring
at the mother cell-bud neck required for cytokinesis and that is
supposed to act as a scaffold for many proteins involved in cell
division.
For Chs2p and Chs1p there is much less information. A
portion of both has been found in a chitosome fraction (42,
50). The bulk of these proteins, however, is in the plasma
membrane (33). It is interesting that none of the Chs proteins
has a signal sequence, although all three use a secretory pathway to reach the membrane.
The regulation of Chs2p is different from that of the other
Chs proteins in that its concentration in the cell changes
greatly during the cell cycle, in contrast to the constant level of
Chs1p and Chs3p (42, 51). The protein has been located by
immunofluorescence at the neck between the mother cell and a
large bud (42). This is the time when presumably the zymogen
form of the enzyme is activated and construction of the primary
septum proceeds (Fig. 2C). Immediately thereafter, Chs2p is
transported to the vacuole and destroyed (42). A concomitant
sharp decrease in its level was detected (42, 51).