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PubMed2000

Corticotropin releasing hormone and related peptides can act as bioregulatory factors in human keratinocytes.

Following previous findings in human skin of the functional expression of genes for the corticotropin releasing hormone (CRH) receptor type 1 (CRH-R1) and CRH itself, we searched for local phenotypic effects for peptides related to CRH. We now report that CRH, sauvagine, and urocortin inhibit proliferation of human HaCaT keratinocytes in a dose-dependent manner. The peptides produced variable cyclic adenosine 3':5'-monophosphate stimulation, with CRH having the highest potency. Binding of iodine 125 CRH to intact keratinocytes was inhibited by increasing doses of CRH, sauvagine, or urocortin, all showing equal inhibitory potency. Immunocytochemistry identified CRH-R1 immunoreactivity in HaCaT keratinocytes. In conclusion, CRH (exogenous or produced locally) and the related urocortin and sauvagine peptides can modify human keratinocyte phenotype through a receptor-mediated pathway.

Amphibian Proteins↗

PubMed2002

Splicing regulation as a potential genetic modifier.

Inherited diseases are associated with profound phenotypic variability, which is affected strongly by genetic modifiers. The splicing machinery could be one such modifying system, through a mechanism involving splicing motifs and their interaction with a complex repertoire of splicing factors. Mutations in splicing motifs and changes in levels of splicing factors can result in different splicing patterns. Changes in the level of normal transcripts or in the relative pattern of different mRNA isoforms affect disease expression, leading to phenotypic variability. Here, we discuss the splicing machinery in terms of its significance in disease severity and its potential role as a genetic modifier.

Alternative Splicing↗

PubMed2002

Vestibular dysfunction in the epistatic circler mouse is caused by phenotypic interaction of one recessive gene and three modifier genes.

Vestibular dysfunction is a frequent clinical problem, leading to dizziness and imbalance. Genes play an important role in its etiology, but the genetics are complex and poorly understood. In this study we have analyzed the complex inheritance pattern in the Epistatic circler mouse, which shows circling behavior indicative of vestibular dysfunction in the mouse. This phenotype exists in a proportion of the F2-generation from an intercross between C57L/J and SWR/J mouse strains. Genetic investigation indicates that the circling behavior is caused by a major recessively inherited gene derived from the SWR/J strain (the Ecs-gene) in combination with at least three different modifier genes derived from C57L/J (the Ecl-genes). Genetic mapping made it possible to localize the Ecs-gene to chromosome 14 and the Ecl-genes to chromosome 3, 4, and 13. This study illustrates the feasibility of identifying genes for multifactorial traits in mice.

Animals↗

PubMed2005

Effects of aerobic exercise and obesity phenotype on abdominal fat reduction in response to weight loss.

OBJECTIVE: To test the effects on abdominal fat reduction of adding aerobic exercise training to a diet program and obesity phenotype in response to weight loss. DESIGN: A prospective clinical trial with a 14-week weight-loss intervention design. SETTING AND PARTICIPANTS: In total, 209 overweight and obese women were assigned to four subgroups depending on type of treatment and the subject's obesity phenotype: diet alone (DA) with intra-abdominal fat (IF) obesity (> or =mean IF area), diet plus exercise (DE) with IF obesity, DA with abdominal subcutaneous fat (ASF) obesity (<mean IF area) and DE with ASF obesity. Abdominal fat areas were evaluated by CT scans, with values adjusted for selected variables. RESULTS: Values were adjusted for age, menopausal status and change in body weight and total fat mass. The IF reductions were significantly (P<0.0001) greater in subjects with IF obesity phenotype (-45.1 cm2) compared to the ASF obesity phenotype (-22.2 cm2). The ASF reductions were significantly (P<0.001) greater for subjects with ASF obesity (-74.5 cm2) compared to IF obesity (-55.5 cm2). For IF obesity, the IF reduction was significantly (P<0.01) greater in the DE group (-49.3 cm2) than in the DA group (-37.8 cm2). CONCLUSION: These results suggest that for individuals with IF obesity, the efficacy on reducing IF of adding aerobic exercise training to a diet-alone weight-reduction program is more prominent (-49.3 cm2/-37.8 cm2=1.3 times) compared with DA. Moreover, abdominal fat reduction was found to be modified by obesity phenotype in response to weight loss.

Adult↗

PubMed2005

An early step in wobble uridine tRNA modification requires the Elongator complex.

Elongator has been reported to be a histone acetyltransferase complex involved in elongation of RNA polymerase II transcription. In Saccharomyces cerevisiae, mutations in any of the six Elongator protein subunit (ELP1-ELP6) genes or the three killer toxin insensitivity (KTI11-KTI13) genes cause similar pleiotropic phenotypes. By analyzing modified nucleosides in individual tRNA species, we show that the ELP1-ELP6 and KTI11-KTI13 genes are all required for an early step in synthesis of 5-methoxycarbonylmethyl (mcm5) and 5-carbamoylmethyl (ncm5) groups present on uridines at the wobble position in tRNA. Transfer RNA immunoprecipitation experiments showed that the Elp1 and Elp3 proteins specifically coprecipitate a tRNA susceptible to formation of an mcm5 side chain, indicating a direct role of Elongator in tRNA modification. The presence of mcm5U, ncm5U, or derivatives thereof at the wobble position is required for accurate and efficient translation, suggesting that the phenotypes of elp1-elp6 and kti11-kti13 mutants could be caused by a translational defect. Accordingly, a deletion of any ELP1-ELP6 or KTI11-KTI13 gene prevents an ochre suppressor tRNA that normally contains mcm5U from reading ochre stop codons.

Acetyltransferases↗

PubMed1998

Genetic interactions suggest that Danforth's short tail (Sd) is a gain-of-function mutation.

Danforth's short tail (Sd) is a semidominant mutation on mouse chromosome 2 that acts cell autonomously in the notochord and leads to its distintegration, and thus causes severe defects in somite patterning and vertebral column development. The molecular nature of the Sd gene and mutation is unknown, and it is unclear whether Sd is a loss-of-function mutation and the semidominant inheritance of the Sd phenotype is due to haploinsufficiency, or whether Sd represents a gain-of-function mutation in a gene essential for notochord development and maintenance. Here, we report on the genetic interaction between Sd and an insertional mutation called Etl4lacZ, which provides genetic evidence that Sd is a gain-of-function mutation. Etl4lacZ is an enhancer trap insertion, which gives rise to lacZ expression in distinct cell types, including the notochord. In homozygosity, the lacZ insertion leads to abnormal vertebrae in the caudal part of the vertebral column. Etl4lacZ maps approximately 0.75 cM distal to Sd, and in double heterozy gotes modifies the Sd phenotype contrarily, depending on the chromosomal configuration of the Sd and Etl4lacZ mutations: when Etl4lacZ is present on the chromosome wild type for Sd (Sd+/+ Etl4lacZ; trans configuration), the Sd phenotype is enhanced, i.e., vertebral malformations extend to more anterior positions and the vertebral body of the axis is further reduced. Conversely, when Etl4lacZ is present on the same chromosome as Sd (Sd Etl4lacZ/+ +; cis configuration), the Sd phenotype is attenuated, i.e., vertebral malformations are confined to more posterior levels, and the dens axis, which is severely reduced or absent in Sd heterozygotes, is restored. The different effect of the Etl4lacZ insertion on Sd, depending on its presence in trans or cis, suggests a direct interaction of the transgene insertion with the Sd gene. Additionally, the attenuation of the Sd phenotype by Etl4lacZ in cis suggests that Sd is a gain-of-function mutation and lends support to the idea that Etl4lacZ is a new allele of Sd. Dev. Genet. 23:86-96, 1998.

Alleles↗

PubMed2004

Metabolic profiles to define the genome: can we hear the phenotypes?

There is an increased reliance on genetically modified organisms as a functional genomic tool to elucidate the role of genes and their protein products. Despite this, many models do not express the expected phenotype thought to be associated with the gene or protein. There is thus an increased need to further define the phenotype resultant from a genetic modification to understand how the transcriptional or proteomic network may conspire to alter the expected phenotype. This is best typified by the description of the silent phenotype in genetic manipulations of yeast. High-resolution proton nuclear magnetic resonance ((1)H NMR) spectroscopy provides an ideal mechanism for the profiling of metabolites within biofluids, tissue extracts or, with recent advances, intact tissues. These metabolic datasets can be readily mined using a range of pattern recognition techniques, including hierarchical cluster analysis, principal components analysis, partial least squares and neural networks, with the combined approach being termed metabolomics. This review describes the application of NMR-based metabolomics or metabonomics to genetic and chemical interventions in a number of different species, demonstrating the versatility of such an approach, as well as suggesting how it may be integrated with other "omic" technologies.

Animals↗

PubMed1996

Two distinct apolipoprotein B alleles in mice generated by a single 'in-out' targeting.

'In-out' gene targeting using a hypoxanthine phosphoribosyltransferase (HPRT) minigene was applied to generate two new alleles in the gene (Apob) coding for apolipoprotein B (apo B) in murine embryonic stem (ES) cells. Homologous integration of the targeting vector during the 'in step' disrupted the Apob gene leading to an allele encoding apo B81, having a 19% carboxyl-terminal truncation. All six targeted cells obtained had more than one insert at the locus, and the chromosomal target sequence in four of them was changed during the recombination. These results suggest that concatenation of the targeting vector prior to insertion was needed to generate sufficient gene product to yield the HPRT+ phenotype, and that recombination between the concatenated DNA and endogenous DNA was a gene replacement more frequently than a simple insertion. The 'out step' recombination event which occurs between sequences duplicated in the 'in step', was planned to replace the sequences encoding the putative LDL receptor-binding domains of apo B100 with sequences encoding human beta-globin peptides (designated apo B100-beta). 6-Thioguanine (6-TG) resistant colonies were obtained from all the 'in-step' cell lines tested at frequencies of 10(-5) to 10(-4), but the frequency of physical loss of the HPRT sequences accompanied by retention of the modified Apob sequence was variable, indicating that mechanisms other than a simple excision are responsible for the generation of 6-TG resistance. Mice from the 'in-step' produce apo B81 and display characteristics of familial hypobetalipoproteinemia; some homozygotes develop hydrocephaly or exencephaly. Mice from the 'out-step' produce apo B100-beta and secrete lipoprotein particles containing the modified protein; their phenotypic changes are subtle, suggesting the lack of the putative LDL receptor-binding domains is not sufficient to increase the steady-state level of apo B100-beta particles above that of apo B100 particles in control mice.

Alleles↗

PubMed1998

The lurcher mutation and ionotropic glutamate receptors: contributions to programmed neuronal death in vivo.

The recent positional cloning and physiological characterization of the lurcher mutation resulted in the identification of a novel stimulus that results in neurodegeneration. The catastrophic loss of cerebellar Purkinje cells in lurcher heterozygotes has now been strongly associated with a large constitutive inward current which ultimately activates a programmed form of neuronal death. The completely penetrant and focal nature of the lurcher phenotype gives us an opportunity to investigate the manner in which neurons respond to an aberrant signal in the context of the brain parenchyma. Although there is no human genetic disease that is equivalent to the lurcher mutation at this time, its triggering of programmed neuronal death enables us to pose and address questions that are relevant to a large number of human neurological diseases. The advantage of working in a genetically manipulable in vivo mammalian system is evident: we can address questions relating to gene function in the nervous system in a context that is physiological. Classical genetic analyses looking for molecules that suppress or modify the lurcher phenotype are under way and have now been supplemented with two novel techniques developed in our laboratory: biolistic transfection of cerebellar slices and Bacterial Artificial Chromosome modification. The integration of these novel and classical approaches will facilitate the testing of hypotheses, developed during the course of our study of the lurcher mutation, which explore the propagation of abnormal signals and the initiation of programmed neuronal death in neurons.

Amino Acid Sequence↗

PubMed1993

Isolation of a gene expressed during early embryogenesis from the region of 22q11 commonly deleted in DiGeorge syndrome.

DiGeorge syndrome (DGS) is one of several syndromes associated with deletions within the proximal long-arm of chromosome 22. The region of chromosome 22q11 responsible for the haploinsufficiency syndromes (the DiGeorge Critical Region or DGCR) has been mapped using RFLPs, quantitative Southern blotting and FISH. Similar deletions are seen in the velo-cardio-facial syndrome (VCFS) and familial congenital heart defects. It is not known whether the phenotypic spectrum is the result of the hemizygosity of one gene or whether it is a consequence of contiguous genes being deleted. However, the majority of patients have a large (> = 2Mb deletion). In this paper we report the isolation of a gene, lab name T10, encoding a serine/threonine rich protein of unknown function which maps to the commonly deleted region of chromosome 22q11. Studies in the mouse indicate that it maps to MMU16 and is expressed during early embryogenesis. Although not mapping within the shortest region of overlap for DGS/VCFS, and therefore not the major gene involved in DGS, the expression pattern suggests that this gene may be involved in modifying the haploinsufficient phenotype of hemizygous patients.

Amino Acid Sequence↗

PubMed1997

Expression of the potato leafroll virus ORF0 induces viral-disease-like symptoms in transgenic potato plants.

The role of the open reading frame 0 (ORF0) of luteoviruses in the viral infection cycle has not been resolved, although the translation product (p28) of this ORF has been suggested to play a role in host recognition. To investigate the function of the potato leafroll luteovirus (PLRV) p28 protein, transgenic potato plants were produced containing the ORF0. In the lines in which the ORF0 transcripts could be detected by Northern (RNA) analysis, the plants displayed an altered phenotype resembling virus-infected plants. A positive correlation was observed between levels of accumulation of the transgenic transcripts and severity of the phenotypic aberrations observed. In contrast, potato plants transformed with a modified, untranslatable ORF0 sequence were phenotypically indistinguishable from wild-type control plants. These results suggest that the p28 protein is involved in viral symptom expression. Southern blot analysis showed that the transgenic plants that accumulated low levels of ORF0 transcripts detectable only by reverse transcription-polymerase chain reaction, contained methylated ORF0 DNA sequences, indicating down-regulation of the transgene provoked by the putatively unfavorable effects p28 causes in the plant cell.

Amino Acid Sequence↗

PubMed2005

Alterations of renal phenotype and gene expression profiles due to protein overload in NOD-related mouse strains.

BACKGROUND: Despite multiple causes, Chronic Kidney Disease is commonly associated with proteinuria. A previous study on Non Obese Diabetic mice (NOD), which spontaneously develop type 1 diabetes, described histological and gene expression changes incurred by diabetes in the kidney. Because proteinuria is coincident to diabetes, the effects of proteinuria are difficult to distinguish from those of other factors such as hyperglycemia. Proteinuria can nevertheless be induced in mice by peritoneal injection of Bovine Serum Albumin (BSA). To gain more information on the specific effects of proteinuria, this study addresses renal changes in diabetes resistant NOD-related mouse strains (NON and NOD.B10) that were made to develop proteinuria by BSA overload. METHODS: Proteinuria was induced by protein overload on NON and NOD.B10 mouse strains and histology and microarray technology were used to follow the kidney response. The effects of proteinuria were assessed and subsequently compared to changes that were observed in a prior study on NOD diabetic nephropathy. RESULTS: Overload treatment significantly modified the renal phenotype and out of 5760 clones screened, 21 and 7 kidney transcripts were respectively altered in the NON and NOD.B10. Upregulated transcripts encoded signal transduction genes, as well as markers for inflammation (Calmodulin kinase beta). Down-regulated transcripts included FKBP52 which was also down-regulated in diabetic NOD kidney. Comparison of transcripts altered by proteinuria to those altered by diabetes identified mannosidase 2 alpha 1 as being more specifically induced by proteinuria. CONCLUSION: By simulating a component of diabetes, and looking at the global response on mice resistant to the disease, by virtue of a small genetic difference, we were able to identify key factors in disease progression. This suggests the power of this approach in unraveling multifactorial disease processes.

Animals↗

PubMed2006

[X chromosome inactivation patterns in patients with Rett syndrome and their mothers and the parental origin of the priority inactive X chromosome].

OBJECTIVE: Rett syndrome (RTT) is a severe childhood neurodevelopmental disorder mainly affecting females. The pathogenic gene is located at Xq28, which codes for the methyl-CpG-binding protein 2. MECP2 gene is affected by X chromosome inactivation (XCI). The different XCI patterns of females could affect the expression ratios of pathogenic gene, causing changes in clinical symptoms. In order to understand the XCI patterns in RTT patients and the relationship between XCI pattern, genotype and phenotype, the XCI patterns in patients with RTT and their mothers, the parental origin of the priority inactive X chromosome in RTT, and the relations of XCI patterns with genotype and phenotype in RTT cases were analyzed. METHODS: Genomic DNA was extracted from peripheral blood of 55 cases with RTT (52 with MECP2 mutations, 3 without mutations), 53 mothers of RTT cases and 48 normal female controls. DNA was digested with methylation sensitive restriction endonuclease Hpa II. Then the undigested and digested DNAs were amplified via PCR for the first exon of human androgen receptor (AR) gene. PCR products were analyzed by Genescan. RESULTS: The heterozygotic rates of AR gene were 82%, 77% and 83% in RTT patients, mothers and controls, respectively. XCI distribution pattern of RTT was different from that of the mothers and control, P < 0.05. More mothers and controls than RTT patients were in the area of XCI 50:50 - 59:41. The differences between them were statistically significant (P < 0.05). No significant difference in XCI distribution patterns between mothers and the control groups was found (P > 0.05). Non-random XCI rates in the areas of XCI >or= 65:35 and >or= 80:20 were 53.35% and 17.8%, respectively, in RTT patients, compared with the mothers group (36.6%, 7.3%) and control group (35%, 10%), it was higher in RTT patients, but the difference was not statistically significant (P > 0.05). In 18 of 21 cases with XCI >or= 65:35, the priority inactive X chromosome was of paternal origin (85.7%). Variable XCI patterns were observed in the same gene mutation patients. The highly skewed XCI as well as the random XCI were found in patients with mild, severe and typical phenotype. The rate of highly skewed XCI in atypical patients was higher than that in typical RTT patients. The rate of highly skewed XCI in T158M was higher than the other type mutations. No highly skewed XCI was observed in cases with R133C mutation. CONCLUSION: The XCI distribution pattern of RTT patients was different from that of RTT mother and control groups. There was no significant difference in XCI distribution patterns between mothers and the control groups. It was not a main genetic pattern in RTT that mothers as the carriers to transmit the pathogenic gene to the patients. Non-random XCI was not the main XCI pattern in RTT patients. The priority inactive X chromosome was mainly of paternal origin. XCI could modify the clinical phenotype of RTT, but had limitations in explaining all the phenotypes manifested in RTT cases.

Adolescent↗

PubMed2005

Interferon-alpha2a is sufficient for promoting dendritic cell immunogenicity.

Type I interferons (IFNs) are widely used therapeutically. IFN-alpha2a in particular is used as an antiviral agent, but its immunomodulatory properties are poorly understood. Dendritic cells (DCs) are the only antigen-presenting cells able to prime naive T cells and therefore play a crucial role in initiating the adaptive phase of the immune response. We studied the effects of IFN-alpha2a on DC maturation and its role in determining Th1/Th2 equilibrium. We found that IFN-alpha2a induced phenotypic maturation of DCs and increased their allostimulatory capacity. When dendritic cells were stimulated simultaneously by CD40 ligation and IFN-alpha2a, the production of interleukin (IL)-10 and IL-12 was increased. In contrast, lipopolysaccharide (LPS) stimulation in the presence of IFN-alpha2a mainly induced IL-10 release. The production of IFN-gamma and IL-5 by the responder naive T cells was also amplified in response to IFN-alpha2a-treated DCs. Furthermore, IL-12 production by IFN-alpha2a-treated DCs was enhanced further in the presence of anti-IL-10 antibody. Different results were obtained when DCs were treated simultaneously with IFN-alpha2a and other maturation factors, in particular LPS, and then stimulated by CD40 ligation 36 h later. Under these circumstances, IFN-alpha2a did not modify the DC phenotype, and the production of IL-10/IL-12 and IFN-gamma/IL-5 by DCs and by DC-stimulated naive T cells, respectively, was inhibited compared to the effects on DCs treated with maturation factors alone. Altogether, this work suggests that IFN-alpha2a in isolation is sufficient to promote DC activation, however, other concomitant events, such as exposure to LPS during a bacterial infection, can inhibit its effects. These results clarify some of the in vivo findings obtained with IFN-alpha2a and have direct implications for the design of IFN-alpha-based vaccines for immunotherapy.

Animals↗

PubMed1993

Nerve growth factor suppresses the transforming phenotype of human prolactinomas.

The most effective therapy of human prolactinomas is represented by dopamine D-2 receptor agonists; there is, however, a population of nonresponder patients who require surgical intervention. In the present study, we report that prolactinomas totally resistant to pharmacological therapy have a high potential of both growing in soft agar and forming tumors in nude mice and lack D-2 receptors for dopamine. These tumors express the receptors for nerve growth factor (NGF) and are sensitive to its differentiating activity. After exposure to NGF for 4 days, prolactinoma cells decreased their proliferation rate, lost their capability to form colonies in soft agar, lost their tumorigenic activity in nude mice, and reexpressed the lactotroph-specific D-2 receptor protein inhibiting prolactin release. These effects were permanent after NGF withdrawal and were reproducible in vivo in nude mice transplanted with the tumors. NGF in fact remarkably and lastingly depressed tumor growth and induced expression of D-2 receptors when injected intravenously once a day for 5 days into prolactinoma-bearing nude mice. These data suggest that NGF may induce a long-lasting switch of gene expression in human prolactinomas, modifying their transforming phenotype and reverting them to more differentiated, less malignant, dopamine-sensitive lactotroph-like cells. The possibility thus arises that short-term treatment with NGF may restore the refractory patients to conventional pharmacological therapy with D-2 agonists.

Animals↗

PubMed1998

A genetic screen for elements of the network that regulates neurogenesis in Drosophila.

The development of external sensory organs on the notum of Drosophila is promoted by the proneural genes achaete and scute. Their activity defines proneural cell clusters in the wing imaginal disc. Ectopic expression, under control of the GAL4 system, of the proneural gene lethal of scute (l'sc) causes the development of ectopic bristles. Persistent ectopic expression of l'sc is not sufficient to impose a neural fate on any given cell. This implies that mutual inhibition, mediated by the Notch signaling pathway, occurs among the cells of the ectopic proneural cluster. Consequently, the dominant, quantifiable phenotype associated with ectopic expression of l'sc is modified by mutations in genes known to be involved in neurogenesis. This phenotype has been utilized to screen for dominant enhancers and suppressors that modify the number of ectopic bristles. In this way, about 100,000 progeny of EMS or X-ray-treated flies have been analyzed to identify autosomal genes involved in regulation of the neural fate. In addition 1200 chromosomes carrying lethal P-element insertions were screened for modifiers. Besides mutations in genes expected to modify the phenotype, we have isolated mutations in six genes not known so far to be involved in neurogenesis.

Alleles↗

PubMed1994

Increased proliferative activity, loss of beta-adrenergic receptor function and class I major histocompatibility complex antigen surface expression in a modified lymphoma cell line.

The molecular interaction of class I major histocompatibility complex (MHC) antigens (Ag) and of beta-adrenergic receptors was previously demonstrated on lymphocytes. By long-term culturing with high concentration of foetal calf serum, the murine S49 lymphoma cell line was modified (S49m) giving phenotypic alterations in beta-adrenergic receptors and class I Ag expression. S49m cells displayed a reduced number of beta-adrenergic sites that were uncoupled to the adenylate cyclase system. These were unable to respond to beta agonist stimulation, despite the fact that direct activation of Gs could be achieved with aluminium tetrafluoride. Although S49m cells showed normal expression of the thy 1.2 Ag, they displayed no expression of class I Ag of the d haplotype. This was assessed by the evident lack of cytotoxic activity of specific monoclonal antibodies (Mo Ab) and of their binding. When performing IFI staining on permeabilized cells, we found positive staining with anti-class d Ab inside the cell. This loss of expression and activity of beta-adrenoceptors and the internalization of class I Ag were accompanied by a higher rate of proliferation in S49m cells. The possibility that the loss of both molecules would modify the biology of the cell is also discussed.

Adrenergic beta-Antagonists↗

PubMed1993

Dihydrocytochalasin B enhances transforming growth factor-beta-induced reexpression of the differentiated chondrocyte phenotype without stimulation of collagen synthesis.

Rabbit articular chondrocytes were treated with retinoic acid (RA) to eliminate the differentiated phenotype marked by the synthesis of type II collagen and high levels of proteoglycan. Exposure of such cells to transforming growth factor-beta 1 (TGF-beta 1) in secondary culture under serum-free and RA-free, defined conditions led to reexpression of the differentiated phenotype. The microfilament modifying drug, dihydrocytochalasin B (DHCB), enhanced the effectiveness of TGF-beta 1 and produced a threefold stimulation of type II collagen reexpression (measured by 2-D CNBr peptide mapping) at 0.3 ng/ml TGF-beta 1 without altering total collagen synthesis. Type II collagen reexpression was maximal from 1 to 5 ng/ml TGF-beta 1, with or without DHCB. The effect of DHCB on proteoglycan synthesis was maximal at 1 ng/ml TGF-beta 1. At this dose TGF-beta alone produced no increase in 35SO4 incorporation, while simultaneous treatment with DHCB caused a sevenfold stimulation of proteoglycan synthesis. DHCB-independent stimulation proteoglycan reexpression occurred between 5 and 15 ng/ml TGF-beta 1. In contrast, TGF-beta 1-dependent stimulation of proteoglycan synthesis in differentiated chondrocytes in primary monolayer culture was not substantially affected by DHCB. The collagen data suggest that TGF-beta 1 utilizes separate pathways to control phenotypic change and collagen (matrix) synthesis. Microfilament modification by DHCB selectively enhances the effectiveness of the TGF-beta 1-dependent signaling pathway that controls reexpression of the differentiated phenotype.

Actin Cytoskeleton↗