G-CSF

6 Pages·2000·0.144 MB·English

by  Nagata S.

G-CSF
Shigekazu Nagata
*
Department of Genetics, Osaka University Medical School, 2-2 Yamada-oka Suita,
Osaka, 565-0871, Japan
*corresponding author tel: (cid:135)81-6-6879-3310, fax: (cid:135)81-6-6879-3319,
e-mail: [email protected]
DOI: 10.1006/rwcy.2000.09006.
SUMMARY Structure
G-CSF is a 20–25kDa glycoprotein that specifically G-CSF is a 20–25kDa glycoprotein.
regulates the production of neutrophilic G granulo-
cytes as well as enhancing the functional activities of
Main activities and
matureneutrophils.Itisproducedbyactivatedmacro-
phages, endothelial cells, and fibroblasts. G-CSF is pathophysiological roles
widelyusedclinicallyinthetreatmentofpatientswith
neutropenia after cancer chemotherapy.
G-CSF regulates production of neutrophilic granulo-
cytes, and activates mature neutrophils.
BACKGROUND
Discovery GENE AND GENE REGULATION
Accession numbers
G-CSF (granulocyte colony-stimulating factor) was
discoveredinserumfromendotoxin-treatedmiceasa
factor that stimulates neutrophilic colony formation Human G-CSF: M13008 (Nagata et al., 1986a,b;
from bonemarrowcells, and that inducesdifferentia- Souza et al., 1986)
tion of mouse WEHI-3B D(cid:135) cells into neutrophilic Murine G-CSF: M13926 (Tsuchiya et al., 1986)
granulocytes (Nicola et al., 1983). Some human cell
lines such as squamous carcinoma CHU-2 and blad-
Chromosome location
der carcinoma 5637 were found to constitutively pro-
duceG-CSF(Welteetal.,1985;Nomuraetal.,1986),
and its cDNA was cloned from these cell lines Human chromosome 17q21-q22 (Kanda et al., 1987)
(Nagata et al., 1986a,b; Souza et al., 1986). Mouse chromosome 11 (Buchberg et al., 1988)
Subsequently, mouse G-CSF was identified from a
mouse NFSA cDNA library by crosshybridization
Regulatory sites and corresponding
with human cDNA (Tsuchiya et al., 1986).
transcription factors
Alternative names
There are three regulatory sequences (GPE, G-CSF
G-CSF has also been known as colony-stimulating promoter element) in the 300bp upstream from the
factor3(CSF-3),macrophageandgranulocyteinducer transcription initiation site. GPE1 contains cis-
type 1, granulocyte (MGI-1G), granulocyte-macro- elements for NF(cid:20)B and NF-IL6, while GPE2 is a
phage colony-stimulating factor (cid:12) (GM-CSF(cid:12)), and typical cis-element for OCT (octamer) transcription
pluripotentcolony-stimulatingfactor(pluripoietin). factor(NishizawaandNagata,1990;Nishizawaetal.,
936 Shigekazu Nagata
1990). Accordingly, fibroblasts from NF-IL6-defi- Important homologies
cient mice do not produce G-CSF upon stimulation
by IL-6 or TNF.
HumanandmouseG-CSFsare73.6%identicalatthe
amino acid sequence level (Tsuchiya et al., 1986).
There is a significant sequence homology between
Cells and tissues that express
G-CSF and IL-6 (Hirano et al., 1986). The tertiary
the gene structureofhumanG-CSFissimilartothoseofIL-4,
IL-2, and growth hormone although the similarity in
The G-CSF gene is expressed in monocytes, macro- the primary sequence is not significant (Hill et al.,
phages, endothelial cells, and fibroblasts. 1993; Lovejoy et al., 1993; Zink et al., 1994).
Posttranslational modifications
PROTEIN
Human G-CSF is O-glycosylated at Thr133. The
Accession numbers structure of the sugar moiety attached to human G-
CSF is N-acetylneuraminic acid (cid:11)(2-6)[galactose (cid:12)(1-
Human G-CSF: PID g117564; SwissProt P09919 3)] N-acetylgalactosamine (Souza et al., 1986; Oheda
(Nagata et al., 1986a,b; Souza et al., 1986) et al., 1990).
Mouse G-CSF: PID g117565; SwissProt P09920
(Tsuchiya et al., 1986)
CELLULAR SOURCES AND
TISSUE EXPRESSION
Sequence
Cellular sources that produce
See Figure 1.
Monocytes, macrophages, endothelial cells, and
fibroblasts are induced to express G-CSF by various
Description of protein
stimuli (Metcalf and Nicola, 1985; Broudy et al.,
1987; Koeffler et al., 1987; Seelentag et al., 1987;
The N-terminal 30 amino acids serve as a signal
Kaushansky et al., 1988; Lu et al., 1988; Vellenga
sequence for secretion. Cys36 and Cys42, as well as
et al., 1988; Nishizawa and Nagata, 1990). Some
Cys67 and Cys77, are connected by disulfide bonds
carcinoma cells, such as human squamous carcinoma
(Luetal.,1989).Theisoelectricpointoftheproteinis
CHU-2, bladder carcinoma 5637, glioblastoma
5.5–6.1, depending on the degree of sialylation
U87MG,andhepatomaSK-HEP-1celllinesproduce
(Nomura et al., 1986). It is relatively stable to
G-CSF constitutively (Nomura et al., 1986; Tweardy
extremepHlevels(pH2orpH10),temperature(50%
et al., 1987).
loss of the activity at 70(cid:14)C for 30min), and strong
denaturation agents (6 M guanidine hydrochloride,
8M urea, 0.1% SDS) (Nicola et al., 1983). Eliciting and inhibitory stimuli,
including exogenous and
Discussion of crystal structure endogenous modulators
G-CSFhasafour(cid:11)helixbundlestructure(Hilletal., TNF as well as IL-1 stimulate fibroblasts or
1993; Lovejoy et al., 1993; Zink et al., 1994). endothelial cells to produce G-CSF (Broudy et al.,
Figure 1 Amino acid sequence for human G-CSF.
G-CSF 937
1987; Koeffler et al., 1987; Seelentag et al., 1987; also assayed by its ability to induce differentiation of
Kaushansky et al., 1988). Endotoxins stimulate WEHI-3B D(cid:135) or 32D cells into neutrophils.
macrophagestoproduceG-CSF(MetcalfandNicola,
1985; Nishizawa and Nagata, 1990).
IN VIVO BIOLOGICAL
ACTIVITIES OF LIGANDS IN
RECEPTOR UTILIZATION
ANIMAL MODELS
G-CSF has a unique receptor (G-CSF receptor). Normal physiological roles
ThenormalphysiologicalroleofG-CSFistheproduc-
IN VITRO ACTIVITIES
tion of neutrophils.
In vitro findings
Species differences
G-CSF stimulates the colony formation of neutro-
philic granulocytes in semi-solid cultures of bone
G-CSF hasnospecies-specificity betweenhumanand
marrow cells (Nicola et al., 1983). Unlike other CSFs
mouse (Tsuchiya et al., 1986).
such as GM-CSF and IL-3, G-CSF is rather specific
to progenitor cells of neutrophilic granulocytes. G-
CSFstimulatesnotonlyproliferationanddifferentia-
Knockout mouse phenotypes
tionoftheprogenitors,butalsoprolongsthesurvival
ofthematureneutrophilsandenhancesthefunctional
capacity of the mature neutrophils (Kitagawa et al., Mice lacking the G-CSF gene show chronic
1987; Williams et al., 1990; Yuo et al., 1989). Several neutropenia, are deficient in granulocyte and macro-
myeloidleukemiacelllinessuchasmouseNFS60and phageprogenitorcells,andshowimpairedneutrophil
human TF-1 cells proliferate in response to G-CSF mobilization (Lieschke et al., 1994).
(Tsuchiyaetal.,1986;Kitamuraetal.,1989),whereas
some other myeloid cell lines, such as mouse WEHI-
Transgenic overexpression
3BD(cid:135),32D,andL-G,canbeinducedtodifferentiate
into neutrophilic granulocytes by G-CSF (Nicola
et al., 1983; Valtieri et al., 1987; Lee et al., 1991). Long-term exposure of mice to G-CSF in transgenic
mice causes sustained granulocytosis (Chang et al.,
1989).
Regulatory molecules: Inhibitors
and enhancers Pharmacological effects
G-CSF-induced neutrophilic colony formation from Administration of G-CSF into mice stimulates
bonemarrowisinhibitedbyIFN(cid:13),lymphotoxin,and granulopoiesis (Cohen et al., 1987; Tsuchiya et al.,
TNF(cid:11) (Barber et al., 1987). 1987; Welte et al., 1987).
Bioassays used PATHOPHYSIOLOGICAL ROLES
IN NORMAL HUMANS AND
G-CSF can be assayed by its neutrophilic colony-
DISEASE STATES AND
stimulating activity in semi-solid culture of bone
DIAGNOSTIC UTILITY
marrow cells. In this assay, G-CSF has a specific
activity of about 2(cid:2)108units/mg protein (where
Normal levels and effects
50units/mL is the concentration required for half-
maximal stimulation). G-CSF can be assayed by the
MTT method, or [3H]thymidine incorporation into Serumofhealthypersonscontainslessthan30pg/mL
G-CSF-responsive cells such as NFS-60 cells. It is of G-CSF. Its level increases to 50–2000g/mL in
938 Shigekazu Nagata
patients with acute bacterial infections. The G-CSF marrow transplantation (BMT) to be replaced by
levels rise during the neutropenic phase of cyclic peripheral blood stem cell (PBSC) transplants. This
neutropenia (Watari et al., 1989). procedure allows more rapid hematopoietic recovery
than that seen by traditional BMT (Sheridan et al.,
1992).
IN THERAPY G-CSF has also been used to treat patients with
cyclic neutropenia. While it does not alter the cyclic
Preclinical – How does it affect nature of this disease, G-CSF elevates neutrophil
levelsduringthenadirphase,thuspreventingmanyof
disease models in animals?
the symptoms of the disease (Hammond et al., 1989).
Administration of G-CSF protects neutropenic mice
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