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Richard H Gomer

Publications and source records attributed to Richard H Gomer.

At least 19 recordsLinked to original sources
PubMed2006

Bone marrow-derived fibroblast precursors mediate ischemic cardiomyopathy in mice.

We previously described a mouse model of fibrotic ischemia/reperfusion cardiomyopathy (I/RC) arising from daily, brief coronary occlusion. One characteristic of I/RC was the prolonged elevation of monocyte chemoattractant protein 1 (MCP-1), which was obligate to its phenotype and may contribute to the uptake of bloodborne cells. Here we describe in I/RC hearts a population of small spindle-shaped fibroblasts that were highly proliferative and expressed collagen I and alpha-smooth muscle actin (myofibroblast markers), CD34 (a precursor marker), and CD45 (a hematopoietic marker). These cells represented 3% of all nonmyocyte live cells. To confirm the cells' bone marrow origin, chimeric mice were created by the rescue of irradiated C57BL/6 mice with marrow from ROSA26, a congenic line expressing lacZ. I/RC resulted in a large population of spindle-shaped fibroblasts containing lacZ. We postulated that the fibroblast precursors represented a developmental path for a subset of monocytes, whose phenotype we have shown to be influenced by serum amyloid P (SAP). Thus, we administered SAP in vivo, which markedly reduced the number of proliferative spindle-shaped fibroblasts and completely prevented I/RC-induced fibrosis and global ventricular dysfunction. By contrast, SAP did not suppress the inflammation or chemokine expression seen in I/RC. SAP, a member of the pentraxin family, binds to Fcgamma receptors and modifies the pathophysiological function of monocytes. Our data suggest that SAP interferes with assumption of a fibroblast phenotype in a subset of monocytes and that SAP may be an important regulator in the linkage between inflammation and nonadaptive fibrosis in the heart.

Amyloid↗
PubMed2006

A protein in crude cytosol regulates glucose-6-phosphatase activity in crude microsomes to regulate group size in Dictyostelium.

Dictyostelium discoideum form groups of approximately 2 x 10(4) cells. The group size is regulated in part by a negative feedback pathway mediated by a secreted multipolypeptide complex called counting factor (CF). The CF signal transduction pathway involves CF-repressing internal glucose levels by increasing the K(m) of glucose-6-phosphatase. Little is known about how this enzyme is regulated. Glucose-6-phosphatase is associated with microsomes in both Dictyostelium and mammals. We find that the activity of glucose-6-phosphatase in crude microsomes from cells with high, normal, or low CF activity had a negative correlation with the amount of CF present in these cell lines. In crude cytosols (supernatants from ultracentrifugation of cell lysates), the glucose-6-phosphatase activity had a positive correlation with CF accumulation. The crude cytosols were further fractionated into a fraction containing molecules greater than 10 kDa (S>10K) and molecules less than 10 KDa (S<10K). S>10K from wild-type cells strongly repressed the activity of glucose-6-phosphatase in wild-type microsomes, whereas S>10K from countin(-) cells (cells with low CF activity) significantly increased the activity of glucose-6-phosphatase in wild-type microsomes by decreasing K(m). The regulatory activities in the wild-type and countin(-) S>10Ks are heat-labile and protease-sensitive, suggesting that they are proteins. S<10K from both wild-type and countin(-) cells did not significantly change glucose-6-phosphatase activity. Together, the data suggest that, as a part of a pathway modulating multicellular group size, CF regulates one or more proteins greater than 10 KDa in crude cytosol that affect microsome-associated glucose-6-phosphatase activity.

Animals↗
PubMed2006

Aggregated IgG inhibits the differentiation of human fibrocytes.

Fibrocytes are fibroblast-like cells, which appear to participate in wound healing and are present in pathological lesions associated with asthma, pulmonary fibrosis, and scleroderma. Fibrocytes differentiate from CD14+ peripheral blood monocytes, and the presence of serum delays this process dramatically. We previously purified the factor in serum, which inhibits fibrocyte differentiation, and identified it as serum amyloid P (SAP). As SAP binds to Fc receptors for immunoglobulin G (IgG; Fc gammaRs), Fc gammaR activation may be an inhibitory signal for fibrocyte differentiation. Fc gammaR are activated by aggregated IgG, and we find aggregated but not monomeric, human IgG inhibits human fibrocyte differentiation. Monoclonal antibodies that bind to Fc gammaRI (CD64) or Fc gammaRII (CD32) also inhibit fibrocyte differentiation. Aggregated IgG lacking Fc domains or aggregated IgA, IgE, or IgM do not inhibit fibrocyte differentiation. Incubation of monocytes with SAP or aggregated IgG inhibited fibrocyte differentiation. Using inhibitors of protein kinase enzymes, we show that Syk- and Src-related tyrosine kinases participate in the inhibition of fibrocyte differentiation. These observations suggest that fibrocyte differentiation can occur in situations where SAP and aggregated IgG levels are low, such as the resolution phase of inflammation.

Antibodies, Monoclonal↗
PubMed2006

A 60-kilodalton protein component of the counting factor complex regulates group size in Dictyostelium discoideum.

Much remains to be understood about how a group of cells or a tissue senses and regulates its size. Dictyostelium discoideum cells sense and regulate the size of groups and fruiting bodies using a secreted 450-kDa complex of proteins called counting factor (CF). Low levels of CF result in large groups, and high levels of CF result in small groups. We previously found three components of CF (D. A. Brock and R. H. Gomer, Genes Dev. 13:1960-1969, 1999; D. A. Brock, R. D. Hatton, D.-V. Giurgiutiu, B. Scott, R. Ammann, and R. H. Gomer, Development 129:3657-3668, 2002; and D. A. Brock, R. D. Hatton, D.-V. Giurgiutiu, B. Scott, W. Jang, R. Ammann, and R. H. Gomer, Eukaryot. Cell 2:788-797, 2003). We describe here a fourth component, CF60. CF60 has similarity to acid phosphatases, although it has very little, if any, acid phosphatase activity. CF60 is secreted by starving cells and is lost from the 450-kDa CF when a different CF component, CF50, is absent. Although we were unable to obtain cells lacking CF60, decreasing CF60 levels by antisense resulted in large groups, and overexpressing CF60 resulted in small groups. When added to wild-type cells, conditioned starvation medium from CF60 overexpressor cells as well as recombinant CF60 caused the formation of small groups. The ability of recombinant CF60 to decrease group size did not require the presence of the CF component CF45-1 or countin but did require the presence of CF50. Recombinant CF60 does not have acid phosphatase activity, indicating that the CF60 bioactivity is not due to a phosphatase activity. Together, the data suggest that CF60 is a component of CF, and thus this secreted signal has four different protein components.

Acid Phosphatase↗
PubMed2006

Two circadian timing circuits in Neurospora crassa cells share components and regulate distinct rhythmic processes.

In Neurospora crassa, FRQ, WC-1, and WC-2 proteins comprise the core circadian FRQ-based oscillator that is directly responsive to light and drives daily rhythms in spore development and gene expression. However, physiological and biochemical studies have demonstrated the existence of additional oscillators in the cell that function in the absence of FRQ (collectively termed FRQ-less oscillators [FLOs]). Whether or not these represent temperature-compensated, entrainable circadian oscillators is not known. The authors previously identified an evening-peaking gene, W06H2 (now called clock-controlled gene 16 [ccg-16]), which is expressed with a robust daily rhythm in cells that lack FRQ protein, suggesting that ccg-16 is regulated by a FLO. In this study, the authors provide evidence that the FLO driving ccg-16 rhythmicity is a circadian oscillator. They find that ccg-16 rhythms are generated by a temperature-responsive, temperature-compensated circadian FLO that, similar to the FRQ-based oscillator, requires functional WC-1 and WC-2 proteins for activity. They also find that FRQ is not essential for rhythmic WC-1 protein levels, raising the possibility that this WCFLO is involved in the generation of WC-1 rhythms. The results are consistent with the presence of 2 circadian oscillators within Neurospora cells, which the authors speculate may interact with each other through the shared WC proteins.

Biological Clocks↗
PubMed2005

Mathematically modelling the effects of counting factor in Dictyostelium discoideum.

Size regulation is a crucial feature in many biological systems, with misregulation leading to dysplasia or hyperplasia. The recent discovery of counting factor (CF) in Dictyostelium discoideum will lead to a greater understanding of how the system regulates the size of a group of cells. In this paper we mathematically model the known effects of CF using two different models: a cellular automata model and a discrete continuum hybrid model. With the use of these models we are able to understand how modulation of adhesion and motile forces by CF can facilitate stream breakup. In addition, the modelling suggests a new possible mechanism for stream breakup involving the frequency of cell reorientation.

Animals↗
PubMed2005

A secreted factor represses cell proliferation in Dictyostelium.

Many cells appear to secrete factors called chalones that limit their proliferation, but in most cases the factors have not been identified. We found that growing Dictyostelium cells secrete a 60 kDa protein called AprA for autocrine proliferation repressor. AprA has similarity to putative bacterial proteins of unknown function. Compared with wild-type cells, aprA-null cells proliferate faster, while AprA overexpressing cells proliferate slower. Growing wild-type cells secrete a factor that inhibits the proliferation of wild-type and aprA- cells; this activity is not secreted by aprA- cells. AprA purified by immunoprecipitation also slows the proliferation of wild-type and aprA- cells. Compared with wild type, there is a higher percentage of multinucleate cells in the aprA- population, and when starved, aprA- cells form abnormal structures that contain fewer spores. AprA may thus decrease the number of multinucleate cells and increase spore production. Together, the data suggest that AprA functions as part of a Dictyostelium chalone.

Amino Acid Sequence↗
PubMed2005

Exposure of cells to a cell number-counting factor decreases the activity of glucose-6-phosphatase to decrease intracellular glucose levels in Dictyostelium discoideum.

The development of Dictyostelium discoideum is a model for tissue size regulation, as these cells form groups of approximately 2 x 10(4) cells. The group size is regulated in part by a negative feedback pathway mediated by a secreted multipolypeptide complex called counting factor (CF). CF signal transduction involves decreasing intracellular CF glucose levels. A component of CF, countin, has the bioactivity of the entire CF complex, and an 8-min exposure of cells to recombinant countin decreases intracellular glucose levels. To understand how CF regulates intracellular glucose, we examined the effect of CF on enzymes involved in glucose metabolism. Exposure of cells to CF has little effect on amylase or glycogen phosphorylase, enzymes involved in glucose production from glycogen. Glucokinase activity (the first specific step of glycolysis) is inhibited by high levels of CF but is not affected by an 8-min exposure to countin. The second enzyme specific for glycolysis, phosphofructokinase, is not regulated by CF. There are two corresponding enzymes in the gluconeogenesis pathway, fructose-1,6-bisphosphatase and glucose-6-phosphatase. The first is not regulated by CF or countin, whereas glucose-6-phosphatase is regulated by both CF and an 8-min exposure to countin. The countin-induced changes in the Km and Vmax of glucose-6-phosphatase cause a decrease in glucose production that can account for the countin-induced decrease in intracellular glucose levels. It thus appears that part of the CF signal transduction pathway involves inhibiting the activity of glucose-6-phosphatase, decreasing intracellular glucose levels and affecting the levels of other metabolites, to regulate group size.

Animals↗
PubMed2005

A cysteine-rich extracellular protein containing a PA14 domain mediates quorum sensing in Dictyostelium discoideum.

Much remains to be understood about quorum-sensing factors that allow cells to sense their local density. Dictyostelium discoideum is a simple eukaryote that grows as single-celled amoebae and switches to multicellular development when food becomes limited. As the growing cells reach a high density, they begin expressing discoidin genes. The cells secrete an unknown factor, and at high cell densities the concomitant high levels of the factor induce discoidin expression. We report here the enrichment of discoidin-inducing complex (DIC), an approximately 400-kDa protein complex that induces discoidin expression during growth and development. Two proteins in the DIC preparation, DicA1 and DicB, were identified by sequencing proteolytic digests. DicA1 and DicB were expressed in Escherichia coli and tested for their ability to induce discoidin during growth and development. Recombinant DicB was unable to induce discoidin expression, while recombinant DicA1 was able to induce discoidin expression. This suggests that DicA1 is an active component of DIC and indicates that posttranslational modification is dispensable for activity. DicA1 mRNA is expressed in vegetative and developing cells. The mature secreted form of DicA1 has a molecular mass of 80 kDa and has a 24-amino-acid cysteine-rich repeat that is similar to repeats in Dictyostelium proteins, such as the extracellular matrix protein ecmB/PstA, the prespore cell-inducing factor PSI, and the cyclic AMP phosphodiesterase inhibitor PDI. Together, the data suggest that DicA1 is a component of a secreted quorum-sensing signal regulating discoidin gene expression during Dictyostelium growth and development.

Amino Acid Sequence↗
PubMed2004

[How to reach the right size?].

Very little is known about how the size of an organism, or a specific tissue in an organism, is regulated. Coordinating and regulating the size of tissues is necessary for proper development, wound healing, and regeneration. Defects in a tissue-size regulation mechanism could lead to birth defects or cancer. In addition, there is a strong psychological aspect to some areas of tissue size regulation, as many cosmetic surgery procedures involve enlarging or reducing the size of some body parts. This review addresses the little bit that we know about size regulation. A key concept is that the size of a tissue is the size of the component cells multiplied by the number of those cells. This breaks the size regulation problem down to two parts. The size of cells can be regulated by nutrient sensing and secreted factors, and may have an upper limit due to an upper limit of a genome's ability to produce mRNA's and thus proteins. To regulate the number of cells in a tissue, there are several simple theoretical models involving secreted factors. In one case, the cells can secrete a characteristic factor and the concentration of the factor will increase with the number of cells secreting it, allowing the tissue to sense its own size. In another scenario, a specific cell secretes a limited amount of a factor necessary for the survival of a target population, and this then limits the size of the target population. There are currently several examples of secreted factors that regulate tissue size, including myostatin, which regulates the amount of muscles, leptin, which regulates adipose tissue, and growth hormone and insulin-like growth factors which regulate total mass. In addition, there are factors such as the < > found in Dictyostelium that regulate the breakup of a tissue into sub-groups. A better understanding of how these factors regulate size will hopefully allow us to develop new therapeutic procedures to treat birth defects or diseases that affect tissue size.

Animals↗
PubMed2004

A cell number counting factor regulates Akt/protein kinase B to regulate Dictyostelium discoideum group size.

Little is known about how individual cells can organize themselves to form structures of a given size. During development, Dictyostelium discoideum aggregates in dendritic streams and forms groups of approximately 20,000 cells. D. discoideum regulates group size by secreting and simultaneously sensing a multiprotein complex called counting factor (CF). If there are too many cells in a stream, the associated high concentration of CF will decrease cell-cell adhesion and increase cell motility, causing aggregation streams to break up. The pulses of cyclic AMP (cAMP) that mediate aggregation cause a transient translocation of Akt/protein kinase B (Akt/PKB) to the leading edge of the plasma membrane and a concomitant activation of the kinase activity, which in turn stimulates motility. We found that countin- cells (which lack bioactive CF) and wild-type cells starved in the presence of anticountin antibodies (which block CF activity) showed a decreased level of cAMP-stimulated Akt/PKB membrane translocation and kinase activity compared to parental wild-type cells. Recombinant countin has the bioactivity of CF, and a 1-min treatment of cells with recombinant countin potentiated Akt/PKB translocation to membranes and Akt/PKB activity. Western blotting of total cell lysates indicated that countin does not affect the total level of Akt/PKB. Fluorescence microscopy of cells expressing an Akt/PKB pleckstrin homology domain-green fluorescent protein (PH-GFP) fusion protein indicated that recombinant countin and anti-countin antibodies do not obviously alter the distribution of Akt/PKB PH-GFP when it translocates to the membrane. Our data indicate that CF increases motility by potentiating the cAMP-stimulated activation and translocation of Akt/PKB.

Animals↗
PubMed2003

Inhibition of fibrocyte differentiation by serum amyloid P.

Wound healing and the dysregulated events leading to fibrosis both involve the proliferation and differentiation of fibroblasts and the deposition of extracellular matrix. Whether these fibroblasts are locally derived or from a circulating precursor population is unclear. Fibrocytes are a distinct population of fibroblast-like cells derived from peripheral blood monocytes that enter sites of tissue injury to promote angiogenesis and wound healing. We have found that CD14(+) peripheral blood monocytes cultured in the absence of serum or plasma differentiate into fibrocytes within 72 h. We purified the factor in serum and plasma that prevents the rapid appearance of fibrocytes, and identified it as serum amyloid P (SAP). Purified SAP inhibits fibrocyte differentiation at levels similar to those found in plasma, while depleting SAP reduces the ability of plasma to inhibit fibrocyte differentiation. Compared with sera from healthy individuals and patients with rheumatoid arthritis, sera from patients with scleroderma and mixed connective tissue disease, two systemic fibrotic diseases, were less able to inhibit fibrocyte differentiation in vitro and had correspondingly lower serum levels of SAP. These results suggest that low levels of SAP may thus augment pathological processes leading to fibrosis. These data also suggest mechanisms to inhibit fibrosis in chronic inflammatory conditions, or conversely to promote wound healing.

Adult↗
PubMed2003

Multiple oscillators regulate circadian gene expression in Neurospora.

High-density microarrays were used to profile circadian gene expression in Neurospora crassa cultures grown in constant darkness. We identified 145 clock-controlled genes (ccgs). The ccgs peaked in mRNA accumulation at all phases of the day, with the majority peaking in the late night to early morning. The predicted or known functions of the ccgs demonstrate that the clock contributes to a wide range of cellular processes, including cell signaling, development, metabolism, and stress responses. Although the period of rhythm of most of the ccgs was found to depend on the well characterized frequency (FRQ)-based oscillator, three ccgs appeared to have a rhythm that was significantly short in the long period (29-h) frq7 mutant strain. These ccgs accumulate mRNA rhythmically with a circadian period in a frq-null strain, confirming the existence of a second oscillator in N. crassa.

Circadian Rhythm↗
PubMed2003

Two components of a secreted cell number-counting factor bind to cells and have opposing effects on cAMP signal transduction in Dictyostelium.

A secreted 450-kDa complex of proteins called counting factor (CF) is part of a negative feedback loop that regulates the size of the groups formed by developing Dictyostelium cells. Two components of CF are countin and CF50. Both recombinant countin and recombinant CF50 decrease group size in Dictyostelium. countin- cells have a decreased cAMP-stimulated cAMP pulse, whereas recombinant countin potentiates the cAMP pulse. We find that CF50 cells have an increased cAMP pulse, whereas recombinant CF50 decreases the cAMP pulse, suggesting that countin and CF50 have opposite effects on cAMP signal transduction. In addition, countin and CF50 have opposite effects on cAMP-stimulated Erk2 activation. However, like recombinant countin, recombinant CF50 increases cell motility. We previously found that cells bind recombinant countin with a Hill coefficient of approximately 2, a KH of 60 pm, and approximately 53 sites/cell. We find here that cells also bind 125I-recombinant CF50, with a Hill coefficient of approximately 2, a KH of approximately 15 ng/ml (490 pm), and approximately 56 sites/cell. Countin and CF50 require each other's presence to affect group size, but the presence of countin is not necessary for CF50 to bind to cells, and CF50 is not necessary for countin to bind to cells. Our working hypothesis is that a signal transduction pathway activated by countin binding to cells modulates a signal transduction pathway activated by CF50 binding to cells and vice versa and that these two pathways can be distinguished by their effects on cAMP signal transduction.

Animals↗
PubMed2003

Disruption of aldehyde reductase increases group size in dictyostelium.

Developing Dictyostelium cells form structures containing approximately 20,000 cells. The size regulation mechanism involves a secreted counting factor (CF) repressing cytosolic glucose levels. Glucose or a glucose metabolite affects cell-cell adhesion and motility; these in turn affect whether a group stays together, loses cells, or even breaks up. NADPH-coupled aldehyde reductase reduces a wide variety of aldehydes to the corresponding alcohols, including converting glucose to sorbitol. The levels of this enzyme previously appeared to be regulated by CF. We find that disrupting alrA, the gene encoding aldehyde reductase, results in the loss of alrA mRNA and AlrA protein and a decrease in the ability of cell lysates to reduce both glyceraldehyde and glucose in an NADPH-coupled reaction. Counterintuitively, alrA- cells grow normally and have decreased glucose levels compared with parental cells. The alrA- cells form long unbroken streams and huge groups. Expression of AlrA in alrA- cells causes cells to form normal fruiting bodies, indicating that AlrA affects group size. alrA- cells have normal adhesion but a reduced motility, and computer simulations suggest that this could indeed result in the formation of large groups. alrA- cells secrete low levels of countin and CF50, two components of CF, and this could partially account for why alrA- cells form large groups. alrA- cells are responsive to CF and are partially responsive to recombinant countin and CF50, suggesting that disrupting alrA inhibits but does not completely block the CF signal transduction pathway. Gas chromatography/mass spectroscopy indicates that the concentrations of several metabolites are altered in alrA- cells, suggesting that the Dictyostelium aldehyde reductase affects several metabolic pathways in addition to converting glucose to sorbitol. Together, our data suggest that disrupting alrA affects CF secretion, causes many effects on cellular metabolism, and has a major effect on group size.

Aldehyde Reductase↗
PubMed2003

Disrupted function of tumor necrosis factor-alpha-stimulated gene 6 blocks cumulus cell-oocyte complex expansion.

During ovulation, the oocyte and surrounding somatic cumulus cells contained within a specialized, mucoid matrix are released from the ovary. One matrix component, TNF-alpha-stimulated gene 6 (TSG-6), is a hyaluronan binding protein induced in cumulus cells of preovulatory follicles by the LH surge and is decreased in cumulus cells of COX-2 and prostaglandin E2 (PGE2) receptor subtype EP2 null mice that exhibit impaired ovulation and cumulus expansion. To determine if TSG-6 was hormonally induced in cumulus cells in vitro and was functional during the formation of the expanded matrix, we established a cumulus cell-oocyte complex (COC) culture system. This system was used to analyze the effects of FSH, PGE2, EP2 receptor, and selected protein kinase inhibitors on TSG-6 production as well as specific antibodies to the TSG-6 link module on TSG-6 function. We document that TSG-6 message and protein are induced by cAMP/protein kinase A/MAPK signaling pathways and that blocking these cascades prevents expansion and the production of TSG-6. FSH but not PGE2 rescued expansion and production of TSG-6 in the EP2 null COCs, indicating that generation of a cAMP signal is essential. Furthermore, disruption of the functional interactions between TSG-6, inter-alpha trypsin inhibitor, and hyaluronan with specific antibodies severely altered matrix formation and cumulus expansion, as recorded by time-lapse imaging. Collectively, these results indicate that TSG-6 mRNA is induced in cumulus cells in culture by cAMP and that the secreted TSG-6 protein is a key structural component of the mouse COC matrix.

Animals↗
PubMed2003

CF45-1, a secreted protein which participates in Dictyostelium group size regulation.

Developing Dictyostelium cells aggregate to form fruiting bodies containing typically 2 x 10(4) cells. To prevent the formation of an excessively large fruiting body, streams of aggregating cells break up into groups if there are too many cells. The breakup is regulated by a secreted complex of polypeptides called counting factor (CF). Countin and CF50 are two of the components of CF. Disrupting the expression of either of these proteins results in cells secreting very little detectable CF activity, and as a result, aggregation streams remain intact and form large fruiting bodies, which invariably collapse. We find that disrupting the gene encoding a third protein present in crude CF, CF45-1, also results in the formation of large groups when cells are grown with bacteria on agar plates and then starve. However, unlike countin(-) and cf50(-) cells, cf45-1(-) cells sometimes form smaller groups than wild-type cells when the cells are starved on filter pads. The predicted amino acid sequence of CF45-1 has some similarity to that of lysozyme, but recombinant CF45-1 has no detectable lysozyme activity. In the exudates from starved cells, CF45-1 is present in a approximately 450-kDa fraction that also contains countin and CF50, suggesting that it is part of a complex. Recombinant CF45-1 decreases group size in colonies of cf45-1(-) cells with a 50% effective concentration (EC(50)) of approximately 8 ng/ml and in colonies of wild-type and cf50(-) cells with an EC(50) of approximately 40 ng/ml. Like countin(-) and cf50(-) cells, cf45-1(-) cells have high levels of cytosolic glucose, high cell-cell adhesion, and low cell motility. Together, the data suggest that CF45-1 participates in group size regulation in Dictyostelium.

Amino Acid Sequence↗