BMC Dev
Biol. 2006 Oct 4; Activated
AKT/PKB signaling in C. elegans uncouples
temporally distinct outputs of DAF-2/insulin-like signaling. ·
Gami MS, Iser WB Hanselman KB, Wolkow CA. Laboratory
of Neurosciences, NIA, NIH, BACKGROUND:
In the nematode, Caenorhabditis elegans, a conserved insulin-like signaling pathway
controls larval development, stress resistance and adult lifespan. AGE-1, a
homolog of the p110 catalytic subunit of phosphoinositide
3-kinases (PI3K) comprises the major known effector
pathway downstream of the insulin receptor, DAF-2. Phospholipid
products of AGE-1/PI3K activate AKT/PKB kinase
signaling via PDK-1. AKT/PKB signaling antagonizes nuclear translocation of
the DAF-16/FOXO transcription factor. Reduced AGE-1/PI3K signaling permits
DAF-16 to direct dauer larval arrest and promote
long lifespan in adult animals. In order to study the downstream effectors
of AGE-1/PI3K signaling in C. elegans, we
conducted a genetic screen for mutations that suppress the constitutive dauer arrest phenotype of age-1(mg109) animals.
RESULTS: This report describes mutations recovered in a screen for
suppressors of the constitutive dauer arrest (daf-C) phenotype of age-1(mg109). Two mutations
corresponded to alleles of daf-16. Two mutations were gain-of-function
alleles in the genes, akt-1 and pdk-1, encoding phosphoinositide-dependent
serine/threonine kinases.
A fifth mutation, mg227, located on chromosome X, did not correspond to any
known dauer genes, suggesting that mg227 may
represent a new component of the insulin pathway. Genetic epistasis analysis by RNAi
showed that reproductive development in age-1(mg109);akt-1(mg247)
animals was dependent on the presence of pdk-1. Similarly, reproductive
development in age-1(mg109);pdk-1(mg261) animals
was dependent on akt-1. However, reproductive development in age-1(mg109);
mg227 animals required only akt-1, and pdk-1 activity was dispensable in
this background. Interestingly, while mg227 suppressed dauer
arrest in age-1(mg109) animals, it enhanced the long lifespan phenotype. In
contrast, akt-1(mg247) and pdk-1(mg261) did not affect lifespan or stress
resistance, while both daf-16 alleles fully suppressed these phenotypes.
CONCLUSION: A screen for suppressors of PI3K mutant phenotypes identified
activating mutations in two known pathway components, providing insights
into their regulation. In particular, the interdependence of akt-1 and
pdk-1, even in activated forms, supports the existence of AGE-1-independent
pathways for these phospholipid-dependent kinases. Phenotypic analysis of these alleles shows
that the larval and adult outputs of AGE-1/PI3K are fully separable in
these mutants. PMID: 17020605
[PubMed - indexed for MEDLINE] Curr Biol. 2006 Oct 24;16(20):1977-85. Comment in: Akt and FOXO dysregulation contribute to infection-induced wasting
in Drosophila. Dionne MS, Pham LN, Shirasu-Hiza M, Schneider DS. Department
of Microbiology and Immunology, BACKGROUND:
Studies in Drosophila have taught us a great deal about how animals
regulate the immediate innate immune response, but we still know little
about how infections cause pathology. Here, we examine the pathogenesis
associated with Mycobacterium marinum infection
in the fly. M. marinum is closely related to M.
tuberculosis, which causes tuberculosis in people. RESULTS: A microarray analysis showed that metabolism is
profoundly affected in M. marinum-infected flies.
A genetic screen identified foxo mutants as slower-dying
after infection than wild-type flies. FOXO activity is inhibited by the
insulin effector kinase
Akt; we show that Akt
activation is systemically reduced as a result of M. marinum
infection. Finally, we show that flies infected with Mycobacterium marinum undergo a process like wasting: They
progressively lose metabolic stores, in the form of fat and glycogen. They
also become hyperglycemic. In contrast, foxo
mutants exhibit less wasting. CONCLUSIONS: In people, many
infections--including tuberculosis--can cause wasting, much as we see in
Drosophila. Our study is the first examination of the metabolic
consequences of infection in a genetically tractable invertebrate and gives
insight into the metabolic consequences of mycobacterial
infection, implicating impaired insulin signaling as a key mediator of
these events. These results suggest that the fly can be used to study more
than the immediate innate immune response to infection; it can also be used
to understand the physiological consequences of infection and the immune
response. PMID: 17055976
[PubMed - indexed for MEDLINE] EMBO J. 2006
Dec 21; [Epub ahead of print] Foxo and Fos regulate the decision
between cell death and survival in response to UV irradiation. ·
Luo X, Puig O, Hyun J, Bohmann D, Jasper H. Department
of Biology, Cells
damaged by environmental insults have to be repaired or eliminated to
ensure tissue homeostasis in metazoans. Recent studies suggest that the
balance between cell survival signals and pro-apoptotic stimuli controls
the decision between cell repair and death. How these competing signals are
integrated and interpreted to achieve accurate control over cell fate in
vivo is incompletely understood. Here, we show that the Forkhead
Box O transcription factor Foxo and the AP-1
transcription factor DFos are required downstream
of Jun-N-terminal kinase signaling for the
apoptotic response to UV-induced DNA damage in the developing Drosophila
retina. Both transcription factors regulate the pro-apoptotic gene hid. Our
results indicate that UV-induced apoptosis is repressed by receptor
tyrosine kinase-mediated inactivation of Foxo. These data suggest that integrating stress and
survival signals through Foxo drives the decision
between cell death and repair of damaged cells in vivo. PMID: 17183370
[PubMed - as supplied by publisher Chronic Immune Therapy Induces a Progressive Increase in Intratumoral T Suppressor Activity and a Concurrent
Loss of Tumor-Specific CD8+ T Effectors in her-2/neu Transgenic Mice
Bearing Advanced Spontaneous Tumors. Lab Invest.
2006 May 15; [Epub ahead of print] J
Bone Miner Res. 2006 Jun;21(6):946-55.
Links 
Links
Nair
RE, Kilinc MO, Jones
SA, Egilmez NK.
A single intratumoral injection of IL-12 and
GM-CSF-encapsulated microspheres induces the
complete regression of advanced spontaneous tumors in her-2/neu transgenic
mice. However, tumor regression in this model is transient and long-term
cure is not achieved due to recurrence. Posttherapy
molecular analysis of immune activation/suppression markers within the
tumor microenvironment demonstrated a dramatic up-regulation of IFN-gamma
and a concomitant down-regulation of Forkhead/winged-helix
protein 3 (Foxp3), TGFbeta, and IL-10 expression.
Therapy-induced reversion of immune suppression was transient since all
three markers of suppression recovered rapidly and surpassed pretherapy levels by day 7 after treatment, resulting
in tumor resurgence. Repeated treatment enhanced short-term tumor
regression, but did not augment long-term survival. Serial long-term
analysis demonstrated that although chronic stimulation enhanced the IFN-gamma
response, this was countered by a parallel increase in Foxp3, TGFbeta, and IL-10 expression. Analysis of
tumor-infiltrating T lymphocyte populations showed that the expression of
Foxp3 and IL-10 was associated with CD4(+)CD25(+)
T cells. Repeated treatment resulted in a progressive increase in
tumor-infiltrating CD4(+)CD25(+)Foxp3(+) T
suppressor cells establishing their role in long-term neutralization of antitumor activity. Analysis of tumor-infiltrating CD8(+) T cells demonstrated that although treatment
enhanced IFN-gamma production, antitumor cytotoxicity was diminished. Monitoring of CD8(+) T cells that specifically recognized a dominant
MHC class I her-2/neu peptide showed a dramatic increase in
tetramer-specific CD8(+) T cells after the first treatment; however,
continuous therapy resulted in the loss of this population. These results
demonstrate that both enhanced suppressor activity and deletion of
tumor-specific T cells are responsible for the progressive loss of efficacy
that is associated with chronic immune therapy.
PMID: 16751376 [PubMed - in process]
UBD, a downstream element of FOXP3, allows the
identification of LGALS3, a new marker of human regulatory T cells.
Ocklenburg F, Moharregh-Khiabani D, Geffers R, Janke V, Pfoertner S, Garritsen H, Groebe L, Klempnauer J, Dittmar KE, Weiss
S, Buer J, Probst-Kepper M.
1Junior Research Group for Xenotransplantation,
Department of Visceral and Transplant Surgery,
Here, we report the identification of the ubiquitin-like
gene UBD as a downstream element of FOXP3 in human activated regulatory CD4(+)CD25(hi) T cells (T(reg)).
Retroviral transduction of UBD in human allo-reactive
effector CD4(+) T helper
(T(h)) cells upregulates CD25 and mediates downregulation of IL4 and IL5 expression similar to overexpression of FOXP3. Moreover, UBD impairs T(h) cell proliferation without upregulation
of FOXP3 and impairs calcium mobilization. In the presence of ionomycin, overexpression of
UBD in T(h) cells leads to the induction of IL1R2
that resemble FOXP3-transduced T(h) cells and naturally derived T(reg) cells. A comparison of the transcriptome
of FOXP3- and UBD-transduced T(h)
cells with T(reg) cells allowed the
identification of the gene LGALS3. However, high levels of LGALS3 protein
expression were observed only in human CD4(+)CD25(hi)
derived T(reg) cells and FOXP3-transduced T(h)
cells, whereas little was induced in UBD-transduced
T(h) cells. Thus, UBD contributes to the anergic
phenotype of human regulatory T cells and acts downstream in FOXP3 induced
regulatory signaling pathways, including regulation of LGALS3 expression.
High levels of LGALS3 expression represent a FOXP3-signature of human
antigen-stimulated CD4(+)CD25(hi) derived regulatory T cells.Laboratory
Investigation advance online publication, 15 May 2006;
doi:10.1038/labinvest.3700432.
PMID: 16702978 [PubMed - as supplied by
publisher]
Fibroblast growth factor-2 is an
immediate-early gene induced by mechanical stress in osteogenic
cells.
Li CF, Hughes-Fulford M.
Laboratory of Cell Growth, Northern California Institute for Research and
Education, San Francisco, California, USA.
Fifteen minutes of physiological MS induces FGF-2 in osteogenic
cells. Here, we show that MS induced proliferation in both MC3T3-E1 and BMOp cells isolated from Fgf2(+/+)
mice; Fgf2(-/-) BMOp cells required exogenous
FGF-2 for a normal proliferation response. The induction of fgf-2 is
mediated by PKA and ERK pathways. INTRODUCTION: Mechanical stress (MS)
induces gene expression and proliferation of osteogenic
MC3T3-E1 cells. We have previously shown that physiological levels of MS in
MC3T3-E1 cells causes extracellular
signal-regulated kinase (ERK)1/2
phosphorylation. Here we evaluate the induction
and importance of fibroblast growth factor-2 (FGF-2) for MS-induced proliferation.
MATERIALS AND METHODS: We characterized the MS induction of fgf-2 using a
15-minute pulse of 120 mustrain and studied the
stability of fgf-2 message using actinomycin D.
Bone marrow stromal cells (BMOp)
isolated from Fgf2(-/-) and Fgf2(+/+) mice were used to study the
importance of FGF-2 in MS-induced proliferation. RESULTS: We found that the
induction of fgf-2 by MS is dependent on both protein kinase
A (PKA) and ERK pathways. MS transiently induces fgf-2 within 30 minutes.
FGF-2 receptor (FGFR2) was also significantly increased within 1 h. All
three isoforms of FGF-2 (24, 22, and 18 kDa) were significantly increased by MS. The
MS-mediated increase of fgf-2 mRNA was caused by new synthesis and not
stabilization. Pretreatment of MC3T3-E1 cells with cycloheximide
showed that the induction of fgf-2 did not require new protein synthesis. Pretreating MC3T3-E1 cells with the mitogen-activated
protein kinase (MAPK)/ERK kinase
1/2 (MEK1/2) inhibitor, U0126, or H-89, a PKA inhibitor, significantly
inhibited the induction of fgf-2, showing that mechanical induction of
fgf-2 is dependent on ERK and PKA signaling pathways. The downstream
consequence of a single 15-minute stress pulse was a 3.5-fold increase in
cell number in MC3T3-E1 compared with growth in nonstressed
control cells. In studies using bone marrow osteoprogenitor
cells (BMOp) isolated from Fgf2(+/+)and Fgf2(-/-)
mice, we found that FGF-2 was necessary for a full proliferative
response to MS. CONCLUSIONS: These studies show that FGF-2 is an immediate-early
gene induced by MS, and its expression is mediated by both the PKA and MAPK
signal transduction pathways. FGF-2 was required for a full proliferative response.
PMID: 16753025 [PubMed - in process]