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PubMed2002

The sequence of regulatory events in the sporulation control network of Physarum polycephalum analysed by time-resolved somatic complementation of mutants.

The developmental decision for sporulation of Physarum polycephalum plasmodia is under sensory control by environmental factors like visible light or heat shock and endogenous signals like glucose starvation. Several hours after perceiving an inductive stimulus, plasmodia become committed to sporulation; thereby, they lose their unlimited replicative potential and execute a developmental program that involves differentiation into various cell types required to form a mature fruiting body. Plasmodia are multinuclear single cells which spontaneously fuse upon physical contact. Fusion of mutant plasmodia and cytoplasmic mixing allows complementation studies to be performed at the functional level. Mutant cells altered in their ability to sporulate in response to phytochrome activation by far-red light were cured by fusion with wild-type or other mutant plasmodia. Phytochrome activation in one plasmodium and subsequent fusion with a non-induced plasmodium revealed that complementation of the two mutations depended on (i) which of two genetically distinct plasmodial cells was stimulated; and (ii) on the delay time elapsed between stimulation and cytoplasmic mixing. Such experiments allow us to determine the kinetics and the causal sequence of the regulatory events tagged by mutation.

Animals↗

PubMed2002

Co-regulation of pathogenesis with dimorphism and phenotypic switching in Candida albicans, a commensal and a pathogen.

Candida albicans, a common fungal pathogen of humans, can colonize in many diverse environments of the host and convert between a harmless commensal and a pathogen. Recent advances indicate that C. albicans uses a common set of conserved pathways to regulate dimorphism, mating and phenotypic switching. Major pathways known to regulate dimorphism include a mitogen-activated protein (MAP) kinase pathway through Cph1, the cAMP-dependent protein kinase pathway via Efg1, and Tup1-mediated repression through Rfg1 and Nrg1. The Cph1-mediated MAP kinase pathway is critical for the mating process, while all three pathways are implicated in the regulation of white-opaque switching. All these developmental pathways regulate the expression of hypha-specific and/or phase-specific genes. A high proportion of hypha-specific genes and phase-specific genes encode proteins that contribute directly or indirectly to pathogenesis and virulence of C. albicans. Therefore, virulence genes are co-regulated with cell morphogenesis. This supports a previous notion that the unique aspects of C. albicans commensalism and pathogenesis may lie in the developmental programs of dimorphism and phenotypic switching.

Candida albicans↗

PubMed2006

Embryonic transcription factors in human breast cancer.

Growing evidence suggests that breast cancer cells often reactivate latent developmental programs in order to efficiently execute the multi-step process of tumorigenesis. This review focuses on key transcriptional regulators of embryonic development that are deregulated in breast cancer and discusses the molecular mechanisms by which these proteins control carcinogenesis. Reminiscent of their function during development, embryonic transcription factors regulate changes in gene expression that promote tumor cell growth, cell survival and motility, as well as a morphogenetic process called epithelial-mesenchymal transition (EMT), which is implicated in both breast metastasis and tumor recurrence. Because of their pivotal roles in breast tumor progression, these factors represent valuable new biomarkers for breast cancer detection as well as promising new targets for anti-invasive drugs.

Breast Neoplasms↗

PubMed2005

Regulation of Msx2 gene expression by steroid hormones in human nonmalignant and malignant breast cancer explants cultured in vitro.

Muscle segment homeobox genes, which regulate developmental programs and are expressed in embryonic and adult tissue, play a role in development of some malignancies. There are no reports on the expression of these families of genes in breast cancer tissue. The aim of this study was to compare expression of Msx2 gene in breast cancer of different genotypes as well as in surrounding nonmalignant tissues. Explants obtained during surgery were divided according to their sex steroid receptor status determined by immunocytochemistry. Four explants obtained from malignant and nonmalignant tissue of each individual patient were incubated in a control medium or with the addition of progesterone (10(-7) M) alone, estradiol 17 beta (10(-5) M) or both. The relative level of Msx2 transcripts was evaluated by a semiquantitative RT-PCR and cell proliferation by Alamar Blue test. Results of RT-PCR analysis showed that the relative expression of Msx2 gene depended on the presence of ER/PR receptors both in nonmalignant and malignant tissues Relative amount of Msx2 mRNA was the highest in surrounding nonmalignant ER+/PR- and ER-/PR+ tissue, whereas in ER-/PR- and ER+/PR+ tissue it was 1.4-1.6-fold lower. Tumorigenesis led to about a twofold decrease in the relative amount of Msx2 mRNA except for ER+/PR+ immunophenotype, where no changes were observed. Addition of estradiol or progesterone to the culture of ER-/PR- type tissue explants did not change significantly the relative amount of Msx2 gene mRNA. An opposite effect was observed in ER+/PR- type of tissue. Addition of estradiol alone, or estradiol and progesterone together to tissue culture explants decreased two to three fold the relative amount of Msx2 gene mRNA in both, malignant and surrounding tissues. Progesterone alone had no effect on Msx2 gene expression in this type of tissue. The most complicated regulation was observed in ER+/PR+ type of tissue. Culture of tissue explants supplemented with estradiol significantly increased the relative amount of Msx2 gene mRNA in the surrounding tissue. Progesterone enhanced the stimulatory effect of estradiol in surrounding tissues but not in the malignant tissue. Increased expression of Msx2 correlated with an increased proliferation in ER-/PR- and ER+/PR+ types, but not in ER+/PR- type of tissues. In conclusion, obtained results provide evidence that estrogen affects Msx2 gene expression. Significant changes in the relative amount of Msx2 gene mRNA and lack of canonical ERE element in 5'-upstream sequence of this gene suggest that regulation takes place indirectly probably by protein-protein interaction. The decrease in the relative amount of Msx2 gene mRNA in ER+/PR- type tumor suggests that progesterone also affects Msx2 gene expression by an indirect mechanism(s).

Breast Neoplasms↗

PubMed1987

Changes in the number of chick ciliary ganglion neuron processes with time in cell culture.

The purpose of this study was to describe the shape of chick ciliary ganglion neurons dissociated from embryonic day 8 or 9 ganglia and maintained in vitro. Most of the neurons were multipolar during the first three days after plating, with an average of 6.0 processes extending directly from the cell body. The neurons became unipolar with time. The remaining primary process accounted for greater than 90% of the total neuritic arbor. This striking change in morphology was not due to the selective loss of multipolar cells, or to an obvious decline in the health of apparently intact cells. The retraction of processes was neither prevented nor promoted by the presence of embryonic muscle cells. Process pruning occurred to the same extent and over the same time course whether the cells were plated on a monolayer of embryonic myotubes or on a layer of lysed fibroblasts. Process retraction is not an inevitable consequence of our culture conditions. Motoneurons dissociated from embryonic spinal cords remained multipolar over the same period of time. We conclude that ciliary ganglion neurons breed true in dissociated cell culture in that the multipolar-unipolar transition reflects their normal, in vivo, developmental program.

Animals↗

PubMed1990

Surface cAMP receptors mediate multiple responses during development in Dictyostelium: evidenced by antisense mutagenesis.

Cell surface cAMP receptors (cARs) have been implicated in multiple aspects of development in Dictyostelium. Antisense mutagenesis has recently provided strong evidence that cARs are necessary for aggregation (Klein et al., 1988. Science (Wash. DC). 241:1467-1472). We show here that the expression of cAR1 antisense mRNA which prevents the appearance of cAR1 antigen also prevents the expression of cAMP-binding activity and blocks multiple cAMP-mediated responses. Chemotactic sensitivity to cAMP was lost as were stimulus-induced cAMP and cGMP production. Furthermore, the expression of developmentally regulated marker genes, dependent on repeated cAMP stimulation, was altered. As a result, the developmental program was severely impaired; most of the cells failed to aggregate and undergo further differentiation.

Cell Line, Transformed↗

PubMed1990

Myosin mRNA accumulation and myofibrillogenesis at the myotendinous junction of stretched muscle fibers.

Myofiber growth and myofibril assembly at the myotendinous junction (MTJ) of stretch-hypertrophied rabbit skeletal muscle was studied by in situ hybridization, immunofluorescence, and electron microscopy. In situ hybridization identified higher levels of myosin heavy chain (MHC) mRNA at the MTJ of fibers stretched for 4 d. Electron microscopy at the MTJ of these lengthening fibers revealed a large cytoplasmic space devoid of myofibrils, but containing polysomes, sarcoplasmic reticulum and T-membranes, mitochondria, Golgi complexes, and nascent filament assemblies. Tallies from electron micrographs indicate that myofibril assembly in stretched fibers followed a set sequence of events. (a) In stretched fiber ends almost the entire sarcolemmal membrane was electron dense but only a portion had attached myofibrils. Vinculin, detected by immunofluorescence, was greatly increased at the MTJ membrane of stretched muscles. (b) Thin filaments were anchored to the sarcolemma at the electron dense sites. (c) Thick filaments associated with these thin filaments in an unregistered manner. (d) Z-bodies splice into thin filaments and subsequently thin and thick filaments fall into sarcomeric register. Thus, the MTJ is a site of mRNA accumulation which sets up regional protein synthesis and myofibril assembly. Stretched muscles also lengthen by the addition of myotubes at their ends. After 6 d of stretch these myotubes make up the majority of fibers at the muscle ends. Essentially all these myotubes repeat the developmental program of primary myotubes and express slow MHC. MHC mRNA distribution in myotubes is disorganized as is the distribution of their myofibrils.

Animals↗

PubMed1990

Structure and expression of genes for a class of cysteine-rich proteins of the cuticle layers of differentiating wool and hair follicles.

The major histological components of the hair follicle are the hair cortex and cuticle. The hair cuticle cells encase and protect the cortex and undergo a different developmental program to that of the cortex. We report the molecular characterization of a set of evolutionarily conserved hair genes which are transcribed in the hair cuticle late in follicle development. Two genes were isolated and characterized, one expressed in the human follicle and one in the sheep follicle. Each gene encodes a small protein of 16 kD, containing greater than 50 cysteine residues, ranging from 31 to 36 mol% cysteine. Their high cysteine content and in vitro expression data identify them as ultra-high-sulfur (UHS) keratin proteins. The predicted proteins are composed almost entirely of cysteine-rich and glycine-rich repeats. Genomic blots reveal that the UHS keratin proteins are encoded by related multigene families in both the human and sheep genomes. Tissue in situ hybridization demonstrates that the expression of both genes is localized to the hair fiber cuticle and occurs at a late stage in fiber morphogenesis.

Amino Acid Sequence↗

PubMed1997

Bcl-2 lies downstream of parathyroid hormone-related peptide in a signaling pathway that regulates chondrocyte maturation during skeletal development.

Parathyroid hormone-related peptide (PTHrP) appears to play a major role in skeletal development. Targeted disruption of the PTHrP gene in mice causes skeletal dysplasia with accelerated chondrocyte maturation (Amizuka, N., H. Warshawsky, J.E. Henderson, D. Goltzman, and A.C. Karaplis. 1994. J. Cell Biol. 126:1611-1623; Karaplis, A.C., A. Luz, J. Glowacki, R.T. Bronson, V.L.J. Tybulewicz, H.M. Kronenberg, and R.C. Mulligan. 1994. Genes Dev. 8: 277-289). A constitutively active mutant PTH/PTHrP receptor has been found in Jansen-type human metaphyseal chondrodysplasia, a disease characterized by delayed skeletal maturation (Schipani, E., K. Kruse, and H. Jüppner. 1995. Science (Wash. DC). 268:98-100). The molecular mechanisms by which PTHrP affects this developmental program remain, however, poorly understood. We report here that PTHrP increases the expression of Bcl-2, a protein that controls programmed cell death in several cell types, in growth plate chondrocytes both in vitro and in vivo, leading to delays in their maturation towards hypertrophy and apoptotic cell death. Consequently, overexpression of PTHrP under the control of the collagen II promoter in transgenic mice resulted in marked delays in skeletal development. As anticipated from these results, deletion of the gene encoding Bcl-2 leads to accelerated maturation of chondrocytes and shortening of long bones. Thus, Bcl-2 lies downstream of PTHrP in a pathway that controls chondrocyte maturation and skeletal development.

Animals↗

PubMed1998

SCAR, a WASP-related protein, isolated as a suppressor of receptor defects in late Dictyostelium development.

G protein-coupled receptors trigger the reorganization of the actin cytoskeleton in many cell types, but the steps in this signal transduction cascade are poorly understood. During Dictyostelium development, extracellular cAMP functions as a chemoattractant and morphogenetic signal that is transduced via a family of G protein-coupled receptors, the cARs. In a strain where the cAR2 receptor gene is disrupted by homologous recombination, the developmental program arrests before tip formation. In a genetic screen for suppressors of this phenotype, a gene encoding a protein related to the Wiskott-Aldrich Syndrome protein was discovered. Loss of this protein, which we call SCAR (suppressor of cAR), restores tip formation and most later development to cAR2(-) strains, and causes a multiple-tip phenotype in a cAR2(+) strain as well as leading to the production of extremely small cells in suspension culture. SCAR-cells have reduced levels of F-actin staining during vegetative growth, and abnormal cell morphology and actin distribution during chemotaxis. Uncharacterized homologues of SCAR have also been identified in humans, mouse, Caenorhabditis elegans, and Drosophila. These data suggest that SCAR may be a conserved negative regulator of G protein-coupled signaling, and that it plays an important role in regulating the actin cytoskeleton.

Amino Acid Sequence↗

PubMed1999

Hyphal death during colony development in Streptomyces antibioticus: morphological evidence for the existence of a process of cell deletion in a multicellular prokaryote.

During the life cycle of the streptomycetes, large numbers of hyphae die; the surviving ones undergo cellular differentiation and appear as chains of spores in the mature colony. Here we report that the hyphae of Streptomyces antibioticus die through an orderly process of internal cell dismantling that permits the doomed hyphae to be eliminated with minimum disruption of the colony architecture. Morphological and biochemical approaches revealed progressive disorganization of the nucleoid substructure, followed by degradation of DNA and cytoplasmic constituents with transient maintenance of plasma membrane integrity. Then the hyphae collapsed and appeared empty of cellular contents but retained an apparently intact cell wall. In addition, hyphal death occurred at specific regions and times during colony development. Analysis of DNA degradation carried out by gel electrophoresis and studies on the presence of dying hyphae within the mycelium carried out by electron microscopy revealed two rounds of hyphal death: in the substrate mycelium during emergence of the aerial hyphae, and in the aerial mycelium during formation of the spores. This suggests that hyphal death in S. antibioticus is somehow included in the developmental program of the organism.

Cell Membrane↗

PubMed2000

Expression profiling in the muscular dystrophies: identification of novel aspects of molecular pathophysiology.

We used expression profiling to define the pathophysiological cascades involved in the progression of two muscular dystrophies with known primary biochemical defects, dystrophin deficiency (Duchenne muscular dystrophy) and alpha-sarcoglycan deficiency (a dystrophin-associated protein). We employed a novel protocol for expression profiling in human tissues using mixed samples of multiple patients and iterative comparisons of duplicate datasets. We found evidence for both incomplete differentiation of patient muscle, and for dedifferentiation of myofibers to alternative lineages with advancing age. One developmentally regulated gene characterized in detail, alpha-cardiac actin, showed abnormal persistent expression after birth in 60% of Duchenne dystrophy myofibers. The majority of myofibers ( approximately 80%) remained strongly positive for this protein throughout the course of the disease. Other developmentally regulated genes that showed widespread overexpression in these muscular dystrophies included embryonic myosin heavy chain, versican, acetylcholine receptor alpha-1, secreted protein, acidic and rich in cysteine/osteonectin, and thrombospondin 4. We hypothesize that the abnormal Ca(2)+ influx in dystrophin- and alpha-sarcoglycan-deficient myofibers leads to altered developmental programming of developing and regenerating myofibers. The finding of upregulation of HLA-DR and factor XIIIa led to the novel identification of activated dendritic cell infiltration in dystrophic muscle; these cells mediate immune responses and likely induce microenvironmental changes in muscle. We also document a general metabolic crisis in dystrophic muscle, with large scale downregulation of nuclear-encoded mitochondrial gene expression. Finally, our expression profiling results show that primary genetic defects can be identified by a reduction in the corresponding RNA.

Actins↗

PubMed2002

PKC alpha regulates the hypertrophic growth of cardiomyocytes through extracellular signal-regulated kinase1/2 (ERK1/2).

Members of the protein kinase C (PKC) isozyme family are important signal transducers in virtually every mammalian cell type. Within the heart, PKC isozymes are thought to participate in a signaling network that programs developmental and pathological cardiomyocyte hypertrophic growth. To investigate the function of PKC signaling in regulating cardiomyocyte growth, adenoviral-mediated gene transfer of wild-type and dominant negative mutants of PKC alpha, beta II, delta, and epsilon (only wild-type zeta) was performed in cultured neonatal rat cardiomyocytes. Overexpression of wild-type PKC alpha, beta II, delta, and epsilon revealed distinct subcellular localizations upon activation suggesting unique functions of each isozyme in cardiomyocytes. Indeed, overexpression of wild-type PKC alpha, but not betaI I, delta, epsilon, or zeta induced hypertrophic growth of cardiomyocytes characterized by increased cell surface area, increased [(3)H]-leucine incorporation, and increased expression of the hypertrophic marker gene atrial natriuretic factor. In contrast, expression of dominant negative PKC alpha, beta II, delta, and epsilon revealed a necessary role for PKC alpha as a mediator of agonist-induced cardiomyocyte hypertrophy, whereas dominant negative PKC epsilon reduced cellular viability. A mechanism whereby PKC alpha might regulate hypertrophy was suggested by the observations that wild-type PKC alpha induced extracellular signal-regulated kinase1/2 (ERK1/2), that dominant negative PKC alpha inhibited PMA-induced ERK1/2 activation, and that dominant negative MEK1 (up-stream of ERK1/2) inhibited wild-type PKC alpha-induced hypertrophic growth. These results implicate PKC alpha as a necessary mediator of cardiomyocyte hypertrophic growth, in part, through a ERK1/2-dependent signaling pathway.

Animals↗

PubMed1970

RNA in cytoplasmic and nuclear fractions of cellular slime mold amebas.

A method is described for the rapid separation of cellular slime mold (Dictyostelium discoideum) cells into nuclear and cytoplasmic fractions. Sucrose density sedimentation profiles of radioactivity from cells that had been grown for long or short periods in the presence of uridine-(3)H indicate very low levels of cross-contamination between the fractions. The nuclear fraction contains few, if any, ribosomes. In exponentially growing cells, at least 80% of the ribosomes were associated in polysomal complexes. No loss of counts from pre-labeled rRNA was observed during 2 generations (24 hr) of logarithmic growth and, within the polysomal complexes, the distributions of the preformed material and of rRNA synthesized during the 2 generations were identical. In stationary phase cells that had entered the developmental program leading to fruiting body construction, the rRNA turned over rapidly so that by the end of development at least 75% of the ribosomes fabricated during exponential growth had disappeared and had been replaced by new ones synthesized during the morphogenetic sequence. The preformed ribosomes disappeared preferentially from the monosomal contingent; the newly synthesized ribosomes appeared exclusively in the polysomal contingent and did not appear as monosomes in appreciable numbers for at least 6 hr. The possible significance of this wholesale replacement of ribosomes is discussed.

Amino Acids↗

PubMed1977

Changes in cell surface and cortical cytoplasmic organization during early embryogenesis in the preimplantation mouse embryo.

Membrane topography and organization of cortical cytoskeletal elements and organelles during early embryogenesis of the mouse have been studied by transmission and scanning electron microscopy with improved cellular preservation. At the four- and early eight-cell stages, blastomeres are round, and scanning electron microscopy shows a uniform distribution of microvilli over the cell surface. At the onset of morphogenesis, a reorganization of the blastomere surface is observed in which microvilli becomes restricted to an apical region and the basal zone of intercellular contact. As the blastomeres spread on each other during compaction, many microvilli remain in the basal region of imminent cell-cell contacts, but few are present where the cells have completed spreading on each other. Microvilli on the surface of these embryos contain linear arrays of microfilaments with lateral cross bridges. Microtubules and mitochondria become localized beneath the apposed cell membranes during compaction. Arrays of cortical microtubules are aligned parallel to regions of apposed membranes. During cytokinesis, microtubules become redistributed in the region of the mitotic spindle, and fewer microvilli are present on most of the cell surface. The cell surface and cortical changes initiated during compaction are the first manifestations of cell polarity in embryogenesis. These and previous findings are interpreted as evidence that cell surface changes associated with trophoblast development appear as early as the eight-cell stage. Our observations suggest that morphogenesis involves the activation of a developmental program which coordinately controls cortical cytoplasmic and cell surface organization.

Animals↗

PubMed1979

Distribution of cell surface saccharides on pancreatic cells. II. Lectin-labeling patterns on mature guinea pig and rat pancreatic cells.

The surface saccharide composition of collagenase-dispersed pancreatic cells from adult guinea pig and rat glands was examined by using eight lectins and their ferritin conjugates: Concanavalin A (ConA); Lens culinaris (LCL); Lotus tetragonolobus (LTL); Ricinus communis agglutinins I and II (RCA I, RCA II); Soybean agglutinin (SBA); Ulex europeus lectin (UEL); and wheat germ agglutinin (WGA). Binding studies of iodinated lectins and lectin-ferritin conjugates both revealed one population of saturable, high-affinity receptor sites on the total cell population (approximately 95% acinar cells). Electron microscopy, however, revealed differences in lectin-ferritin binding to the plasmalemma of acinar, centroacinar, and endocrine cells. Whereas acinar cells bound heavily all lectin conjugates, endocrine and centroacinar cells were densely labeled only by ConA, LCL, WGA, and RCA I, and possessed few receptors for LTL, UEL, and SBA. Endocrine and centroacinar cells could be differentiated from each other by using RCA II, which binds to centroacinar cells but not to endocrine cells. Some RCA II receptors appeared to be glycolipids because they were extracted by ethanol and chloroform-methanol in contrast to WGA receptors which resisted solvent treatment but were partly removed by papain digestion. RCA I receptors were affected by neither treatment. The apparent absence of receptors for SBA on endocrine and centroacinar cells, and for RCA II on endocrine cells, was reversed by neuraminidase digestion, which suggested masking of lectin receptors by sialic acid. The absence of LTL and UEL receptors on endocrine and centroacinar cells was not reversed by neuraminidase. We suggest that the differential lectin-binding patterns observed on acinar, centroacinar, and endocrine cells from the adult pancreas surface-carbohydrate-developmental programs expressed during morphogenesis and cytodifferentiation of the gland.

Animals↗

PubMed1981

Monoclonal antibodies against a specific nonhistone chromosomal protein of Drosophila associated with active genes.

Hybridomas secreting monoclonal antibodies have been produced by fusion of NS-1 mouse myeloma cells with the spleen cells of mice inoculated with a 60-65,000-mol wt fraction of proteins released from Drosophila embryo nuclei treated with DNase I. The antibodies secreted by the hybridomas were examined with polytene chromosomes of formaldehyde-fixed salivary gland squashes by an immunofluorescence assay. Most of the clonal antibodies obtained resulted in specific staining of the chromosomes relative to the cytoplasmic debris. In the case of clone 28, the antibodies showed a preferential association with sites of gene activity, both puffs and loci identified as puffing at some time during the third instar and prepupal period. In larvae that were heat shocked (exposed to 35 degrees C for 15 min before removal and fixation of the glands), the antibodies of clone 28 stained preferentially the induced heat-shock loci while continuing to stain most of the normal set of loci. The antigen for clone 28 was identified as a single protein of approximately 62,000 mol wt by using the antibodies followed by 125I-rabbit anti-mouse Ig to stain nitrocellulose replicas of SDS polyacrylamide gels of total chromosomal proteins. This study demonstrates that monoclonal antibodies can be used successfully in immunofluorescence staining of formaldehyde-fixed polytene chromosomes. The results verify the hypothesis that a specific nonhistone chromosomal protein is preferentially associated with the set of loci that includes both active sites and those scheduled to be active at some time in this developmental program. Such proteins may play a general role in the mechanisms of cell determination and gene activation.

Animals↗

PubMed1993

Reconstitution of the subclass-specific expression of CD4 in thymocytes and peripheral T cells of transgenic mice: identification of a human CD4 enhancer.

During thymic maturation, CD4-CD8-TCR- immature thymocytes differentiate through a CD4+CD8+TCRlo intermediate into two functionally distinct mature T cell subsets: helper T cells expressing CD4 and a major histocompatibility complex (MHC) class II-restricted T cell receptor (TCR), and cytotoxic T cells expressing CD8 and and MHC class I-restricted TCR. The mutually exclusive expression of CD4 and CD8 is maintained in the periphery during expansion of these mature T cell subsets. To elucidate the mechanisms controlling CD4 and CD8 expression on differentiating thymocytes and mature peripheral T cells, we have examined the expression of human CD4 gene constructs in the lymphoid tissues of transgenic mice. Our analyses demonstrate that sequences contained within or closely linked to the human CD4 gene are sufficient to reconstitute the appropriate regulation of human CD4 expression on all thymocyte and mature peripheral T cell subsets. Specifically, appropriate developmental regulation was dependent on two sets of sequences, one contained within a 1.3-kb restriction fragment located 6.5 kb upstream of the human CD4 gene, and the other present within or immediately flanking the gene. Nucleotide sequence analysis identified the 1.3-kb restriction fragment as the likely human homologue of an enhancer found 13 kb upstream of the mouse CD4 transcription initiation site. The human CD4 transgenic mice provide a useful system for the identification and characterization of additional sequence elements that participate in human CD4 gene regulation and for the elucidation of regulatory mechanisms governing the developmental program mediating the maturation of the CD4+ and CD8+ peripheral T cell subsets.

Animals↗