Biomedical subjects

J L de la Pompa

Publications and source records attributed to J L de la Pompa.

At least 19 recordsLinked to original sources

PubMed2001

Localized and transient transcription of Hox genes suggests a link between patterning and the segmentation clock.

During development, Hox gene transcription is activated in presomitic mesoderm with a time sequence that follows the order of the genes along the chromosome. Here, we show that Hoxd1 and other Hox genes display dynamic stripes of expression within presomitic mesoderm. The underlying transcriptional bursts may reflect the mechanism that coordinates Hox gene activation with somitogenesis. This mechanism appears to depend upon Notch signaling, as mice deficient for RBPJk, the effector of the Notch pathway, showed severely reduced Hoxd gene expression in presomitic mesoderm. These results suggest a molecular link between Hox gene activation and the segmentation clock. Such a linkage would efficiently keep in phase the production of novel segments with their morphological specification.

Alleles↗

PubMed2001

p53 accumulation, defective cell proliferation, and early embryonic lethality in mice lacking tsg101.

Functional inactivation of the tumor susceptibility gene tsg101 in NIH 3T3 fibroblasts results in cellular transformation and the ability to form metastatic tumors in nude mice. The N-terminal region of tsg101 protein is structurally similar to the catalytic domain of ubiquitin-conjugating enzymes, suggesting a potential role of tsg101 in ubiquitin-mediated protein degradation. The C-terminal domain of TSG101 can function as a repressor of transcription. To investigate the physiological function of tsg101, we generated a null mutation of the mouse gene by gene targeting. Homozygous tsg101-/- embryos fail to develop past day 6.5 of embryogenesis (E6.5), are reduced in size, and do not form mesoderm. Mutant embryos show a decrease in cellular proliferation in vivo and in vitro but no increase in apoptosis. Although levels of p53 transcripts were not affected in tsg101-/- embryos, p53 protein accumulated dramatically, implying altered posttranscriptional control of p53. In addition, transcription of the p53 effector, cyclin-dependent kinase inhibitor p21(WAF-1/CIP-1), was increased 5- to 10-fold, whereas activation of MDM2 transcription secondary to p53 elevation was not observed. Introduction of a p53 null mutation into tsg101-/- embryos rescued the gastrulation defect and prolonged survival until E8.5. These results demonstrate that tsg101 is essential for the proliferative burst before the onset of gastrulation and establish a functional connection between tsg101 and the p53 pathway in vivo.

Animals↗

PubMed1999

Interaction between Notch signalling and Lunatic fringe during somite boundary formation in the mouse.

BACKGROUND: The process of somitogenesis can be divided into three major events: the prepatterning of the mesoderm; the formation of boundaries between the prospective somites; and the cellular differentiation of the somites. Expression and functional studies have demonstrated the involvement of the murine Notch pathway in somitogenesis, although its precise role in this process is not yet well understood. We examined the effect of mutations in the Notch pathway elements Delta like 1 (Dll1), Notch1 and RBPJkappa on genes expressed in the presomitic mesoderm (PSM) and have defined the spatial relationships of Notch pathway gene expression in this region. RESULTS: We have shown that expression of Notch pathway genes in the PSM overlaps in the region where the boundary between the posterior and anterior halves of two consecutive somites will form. The Dll1, Notch1 and RBPJkappa mutations disrupt the expression of Lunatic fringe (L-fng), Jagged1, Mesp1, Mesp2 and Hes5 in the PSM. Furthermore, expression of EphA4, mCer 1 and uncx4.1, markers for the anterior-posterior subdivisions of the somites, is down-regulated to different extents in Notch pathway mutants, indicating a global alteration of pattern in the PSM. CONCLUSIONS: We propose a model for the mechanism of somite border formation in which the activity of Notch in the PSM is restricted by L-fng to a boundary-forming territory in the posterior half of the prospective somite. In this region, Notch function activates a set of genes that are involved in boundary formation and anterior-posterior somite identity.

Animals↗

PubMed1999

Genetic evidence for functional redundancy of Platelet/Endothelial cell adhesion molecule-1 (PECAM-1): CD31-deficient mice reveal PECAM-1-dependent and PECAM-1-independent functions.

Platelet/endothelial cell adhesion molecule-1 (PECAM-1; CD31), a member of the Ig superfamily, is expressed strongly at endothelial cell-cell junctions, on platelets, and on most leukocytes. CD31 has been postulated to play a role in vasculogenesis and angiogenesis, and has been implicated as a key mediator of the transendothelial migration of leukocytes. To further define the physiologic role of CD31, we used targeted gene disruption of the CD31 gene in embryonic stem cells to generate CD31-deficient mice. CD31-deficient mice (CD31KO) are viable and born at the expected Mendelian frequency, remain healthy, and exhibit no obvious vascular developmental defects. In response to inflammatory challenge, polymorphonuclear leukocytes of CD31KO mice are arrested between the vascular endothelium and the basement membrane of inflammatory site mesenteric microvessels, confirming a role for CD31 in the migration of neutrophils through the subendothelial extracellular matrix. Normal numbers of leukocytes are recovered from inflammatory sites in CD31KO mice, however, suggesting that the defect in leukocyte migration across basal lamina observed in the absence of CD31 may be compensated for by the use of other adhesion molecules, or possibly an increased rate of migration. Homing of T lymphocytes in vivo is normal, and CD31KO mice are able to mount a cutaneous hypersensitivity response normally. In addition, CD31-mediated homophilic adhesion does not appear to play a role in platelet aggregation in vitro. This study provides genetic evidence that CD31 is involved in transbasement membrane migration, but does not play an obligatory role in either vascular development or leukocyte migration.

Animals↗

PubMed1998

High cancer susceptibility and embryonic lethality associated with mutation of the PTEN tumor suppressor gene in mice.

BACKGROUND: Germ-line and sporadic mutations in the tumor suppressor gene PTEN (also known as MMAC or TEP1), which encodes a dual-specificity phosphatase, cause a variety of cancers such as Cowden disease, glioblastoma, endometrial carcinoma and prostatic cancer. PTEN is widely expressed, and Cowden disease consistently affects various organ systems, suggesting that the PTEN protein must have an important, although as yet poorly understood, function in cellular physiology. RESULTS: Homozygous mutant mice lacking exons 3-5 of the PTEN gene (mPTEN3-5) had severely expanded and abnormally patterned cephalic and caudal regions at day 8.5 of gestation. Embryonic death occurred by day 9.5 and was associated with defective chorio-allantoic development. Heterozygous mPTEN3-5 mice had an increased incidence of tumors, especially T-cell lymphomas; gamma-irradiation reduced the time lapse of tumor formation. DNA analysis of these tumors revealed the deletion of the mPTEN gene due to loss of heterozygosity of the wild-type allele. Tumors associated with loss of heterozygosity in mPTEN showed elevated phosphorylation of protein kinase B (PKB, also known as Akt kinase), thus providing a functional connection between mPTEN and a murine proto-oncogene (c-Akt) involved in the development of lymphomas. CONCLUSIONS: The mPTEN gene is fundamental for embryonic development in mice, as mPTEN3-5 mutant embryos died by day 9.5 of gestation, with patterning defects in cephalic and caudal regions and defective placentation. Heterozygous mice developed lymphomas associated with loss of heterozygosity of the wild-type mPTEN allele, and tumor appearance was accelerated by gamma-irradiation. These lymphomas had high levels of activated Akt/PKB, the protein product of a murine proto-oncogene with anti-apoptotic function, associated with thymic lymphomas. This suggests that tumors associated with mPTEN loss of heterozygosity may arise as a consequence of an acquired survival advantage. We provide direct evidence of the role of mPTEN as a tumor suppressor gene in mice, and establish the mPTEN mutant mouse as an experimental model for investigating the role of PTEN in cancer progression.

Animals↗

PubMed1998

Negative regulation of PKB/Akt-dependent cell survival by the tumor suppressor PTEN.

PTEN is a tumor suppressor with sequence homology to protein tyrosine phosphatases and the cytoskeletal protein tensin. mPTEN-mutant mouse embryos display regions of increased proliferation. In contrast, mPTEN-deficient immortalized mouse embryonic fibroblasts exhibit decreased sensitivity to cell death in response to a number of apoptotic stimuli, accompanied by constitutively elevated activity and phosphorylation of protein kinase B/Akt, a crucial regulator of cell survival. Expression of exogenous PTEN in mutant cells restores both their sensitivity to agonist-induced apoptosis and normal pattern of PKB/Akt phosphorylation. Furthermore, PTEN negatively regulates intracellular levels of phosphatidylinositol (3,4,5) trisphosphate in cells and dephosphorylates it in vitro. Our results show that PTEN may exert its role as a tumor suppressor by negatively regulating the PI3'K/PKB/Akt signaling pathway.

Animals↗

PubMed1998

Differential requirement for caspase 9 in apoptotic pathways in vivo.

Mutation of Caspase 9 (Casp9) results in embryonic lethality and defective brain development associated with decreased apoptosis. Casp9-/- embryonic stem cells and embryonic fibroblasts are resistant to several apoptotic stimuli, including UV and gamma irradiation. Casp9-/- thymocytes are also resistant to dexamethasone- and gamma irradiation-induced apoptosis, but are surprisingly sensitive to apoptosis induced by UV irradiation or anti-CD95. Resistance to apoptosis is accompanied by retention of the mitochondrial membrane potential in mutant cells. In addition, cytochrome c is translocated to the cytosol of Casp9-/- ES cells upon UV stimulation, suggesting that Casp9 acts downstream of cytochrome c. Caspase processing is inhibited in Casp9-/- ES cells but not in thymocytes or splenocytes. Comparison of the requirement for Casp9 and Casp3 in different apoptotic settings indicates the existence of at least four different apoptotic pathways in mammalian cells.

Animals↗

PubMed1998

FADD: essential for embryo development and signaling from some, but not all, inducers of apoptosis.

FADD (also known as Mort-1) is a signal transducer downstream of cell death receptor CD95 (also called Fas). CD95, tumor necrosis factor receptor type 1 (TNFR-1), and death receptor 3 (DR3) did not induce apoptosis in FADD-deficient embryonic fibroblasts, whereas DR4, oncogenes E1A and c-myc, and chemotherapeutic agent adriamycin did. Mice with a deletion in the FADD gene did not survive beyond day 11.5 of embryogenesis; these mice showed signs of cardiac failure and abdominal hemorrhage. Chimeric embryos showing a high contribution of FADD null mutant cells to the heart reproduce the phenotype of FADD-deficient mutants. Thus, not only death receptors, but also receptors that couple to developmental programs, may use FADD for signaling.

Adaptor Proteins, Signal Transducing↗

PubMed1998

Role of the NF-ATc transcription factor in morphogenesis of cardiac valves and septum.

In lymphocytes, the expression of early immune response genes is regulated by NF-AT transcription factors which translocate to the nucleus after dephosphorylation by the Ca2+-dependent phosphatase, calcineurin. We report here that mice bearing a disruption in the NF-ATc gene fail to develop normal cardiac valves and septa and die of circulatory failure before day 14.5 of development. NF-ATc is first expressed in the heart at day 7.5, and is restricted to the endocardium, a specialized endothelium that gives rise to the valves and septum. Within the endocardium, specific inductive events appear to activate NF-ATc: it is localized to the nucleus only in endocardial cells that are adjacent to the interface with the cardiac jelly and myocardium, which are thought to give the inductive stimulus to the valve primordia. Treatment of wild-type embryos with FK506, a specific calcineurin inhibitor, prevents nuclear localization of NF-ATc. These data indicate that the Ca2+/calcineurin/NF-ATc signalling pathway is essential for normal cardiac valve and septum morphogenesis; hence, NF-ATc and its regulatory pathways are candidates for genetic defects underlying congenital human heart disease.

Animals↗

PubMed1998

The tumor suppressor gene Smad4/Dpc4 is required for gastrulation and later for anterior development of the mouse embryo.

Mutations in the SMAD4/DPC4 tumor suppressor gene, a key signal transducer in most TGFbeta-related pathways, are involved in 50% of pancreatic cancers. Homozygous Smad4 mutant mice die before day 7.5 of embryogenesis. Mutant embryos have reduced size, fail to gastrulate or express a mesodermal marker, and show abnormal visceral endoderm development. Growth retardation of the Smad4-deficient embryos results from reduced cell proliferation rather than increased apoptosis. Aggregation of mutant Smad4 ES cells with wild-type tetraploid morulae rescues the gastrulation defect. These results indicate that Smad4 is initially required for the differentiation of the visceral endoderm and that the gastrulation defect in the epiblast is secondary and non-cell autonomous. Rescued embryos show severe anterior truncations, indicating a second important role for Smad4 in anterior patterning during embryogenesis.

Alleles↗

PubMed1998

neurogenin1 is essential for the determination of neuronal precursors for proximal cranial sensory ganglia.

The NEUROGENINS (NGNs) are neural-specific basic helix-loop-helix (bHLH) transcription factors. Mouse embryos lacking ngn1 fail to generate the proximal subset of cranial sensory neurons. ngn1 is required for the activation of a cascade of downstream bHLH factors, including NeuroD, MATH3, and NSCL1. ngn1 is expressed by placodal ectodermal cells and acts prior to neuroblast delamination. Moreover, NGN1 positively regulates the Delta homolog DLL1 and can be negatively regulated by Notch signaling. Thus, ngn1 functions similarly to the proneural genes in Drosophila. However, the initial pattern of ngn1 expression appears to be Notch independent. Taken together with the fact that ectopic ngn1 expression can convert ectodermal cells to neurons in Xenopus (Ma et al., 1996), these data and those of Fode et al. (1998 [this issue of Neuron]) identify ngns as vertebrate neuronal determination genes, analogous to myoD and myf5 in myogenesis.

Animals↗

PubMed1998

Developmental studies of Brca1 and Brca2 knock-out mice.

In humans, the inheritance of mutations in the breast cancer susceptibility genes BRCA1 and BRCA2 increases the risk of developing breast and ovarian cancer. To study their biological function and to create animal models for these cancer susceptibility genes, several strains of mice mutated in the homologous genes Brca1 and Brca2 have been generated by gene targeting. Analyses of these "knock-out" mouse mutants have provided invaluable knowledge about the function of these genes. Brca1 and Brca2 null mutants are similar in phenotype: mutations in both genes result in embryonic lethality and the developing embryos show signs of a cellular proliferation defect associated with activation of the p53 pathway. The significance of this activation, as well as the role of these cancer susceptibility genes in DNA damage repair, is discussed.

Animals↗

PubMed1997

Brca2 is required for embryonic cellular proliferation in the mouse.

Mutations of the tumor suppressor gene BRCA2 are associated with predisposition to breast and other cancers. Homozygous mutant mice in which exons 10 and 11 of the Brca2 gene were deleted by gene targeting (Brca2(10-11)) die before day 9.5 of embryogenesis. Mutant phenotypes range from severely developmentally retarded embryos that do not gastrulate to embryos with reduced size that make mesoderm and survive until 8.5 days of development. Although apoptosis is normal, cellular proliferation is impaired in Brca2(10-11) mutants, both in vivo and in vitro. In addition, the expression of the cyclin-dependent kinase inhibitor p21 is increased. Thus, Brca2(10-11) mutants are similar in phenotype to Brca1(5-6) mutants but less severely affected. Expression of either of these two genes was unaffected in mutant embryos of the other. This study shows that Brca2, like Brca1, is required for cellular proliferation during embryogenesis. The similarity in phenotype between Brca1 and Brca2 mutants suggests that these genes may have cooperative roles or convergent functions during embryogenesis.

Animals↗

PubMed1997

Early lethality, functional NF-kappaB activation, and increased sensitivity to TNF-induced cell death in TRAF2-deficient mice.

TRAF2 is an intracellular signal-transducing protein recruited to the TNFR1 and TNFR2 receptors following TNF stimulation. To investigate the physiological role of TRAF2, we generated TRAF2-deficient mice. traf2-/- mice appeared normal at birth but became progressively runted and died prematurely. Atrophy of the thymus and spleen and depletion of B cell precursors also were observed. Thymocytes and other hematopoietic progenitors were highly sensitive to TNF-induced cell death and serum TNF levels were elevated in these TRAF2-deficient animals. Examination of traf2-/- cells revealed a severe reduction in TNF-mediated JNK/SAPK activation but a mild effect on NF-kappaB activation. These results suggest that TRAF2-independent pathways of NF-kappaB activation exist and that TRAF2 is required for an NF-kappaB-independent signal that protects against TNF-induced apoptosis.

Animals↗

PubMed1997

Partial rescue of Brca1 (5-6) early embryonic lethality by p53 or p21 null mutation.

Mutations in the mouse Brca1 gene cause lethality at different embryonic stages. We have shown that Brca1 mutant embryos, in which the fifth and sixth exons of Brca1 are deleted die before E7.5 and show decreased cellular proliferation. Brca1 mutants also show decreased expression of mdm2, a gene encoding an inhibitor of p53 activity. Thus, we have proposed that the reduction in mdm2 expression in Brca1 (5-6) mutants might lead to increased p53 activity. Consistent with this finding, the expression of p21, which encodes a G1 cell cycle inhibitor and is a target for p53 transcriptional activation was dramatically increased in the Brca1 (5-6) mutants, suggesting that impaired cellular proliferation could be due to a G1 cell-cycle arrest, caused by increased p21 levels. To test this hypothesis, we generated mice double mutant for Brca1 (5-6) and p53, or Brca1 (5-6) and p21. Mutation in either p53 or p21 prolonged the survival of Brca1 (5-6) mutant embryos from E7.5 to E9.5. The development of most Brca1 (5-6): p21 double-mutant embryos was comparable to that of their wild-type littermates, although no mutant survived past E10.5. The fact that mutation of neither p53 nor p21 completely rescued Brca1 (5-6) embryos suggests that their lethality is likely due to a multi-factorial process.

Animals↗

PubMed1997

Conservation of the Notch signalling pathway in mammalian neurogenesis.

The Notch pathway functions in multiple cell fate determination processes in invertebrate embryos, including the decision between the neuroblast and epidermoblast lineages in Drosophila. In the mouse, targeted mutation of the Notch pathway genes Notch1 and RBP-Jk has demonstrated a role for these genes in somite segmentation, but a function in neurogenesis and in cell fate decisions has not been shown. Here we show that these mutations lead to altered expression of the Notch signalling pathway homologues Hes-5, Mash-1 and Dll1, resulting in enhanced neurogenesis. Precocious neuronal differentiation is indicated by the expanded expression domains of Math4A, neuroD and NSCL-1. The RBP-Jk mutation has stronger effects on expression of these genes than does the Notch1 mutation, consistent with functional redundancy of Notch genes in neurogenesis. Our results demonstrate conservation of the Notch pathway and its regulatory mechanisms from fly to mouse, and support a role for the murine Notch signalling pathway in the regulation of neural stem cell differentiation.

Animals↗

PubMed1996

The tumor suppressor gene Brca1 is required for embryonic cellular proliferation in the mouse.

Mutations of the BRCA1 gone in humans are associated with predisposition to breast and ovarian cancers. We show here that Brca1+/- mice are normal and fertile and lack tumors by age eleven months. Homozygous Brca1(5-6) mutant mice die before day 7.5 of embryogenesis. Mutant embryos are poorly developed, with no evidence of mesoderm formation. The extraembryonic region is abnormal, but aggregation with wild-type tetraploid embryos does not rescue the lethality. In vivo, mutant embryos do not exhibit increased apoptosis but show reduced cell proliferation accompanied by decreased expression of cyclin E and mdm-2, a regulator of p53 activity. The expression of cyclin-dependent kinase inhibitor p21 is dramatically increased in the mutant embryos. Buttressing these in vivo observations is the fact that mutant blastocyst growth is grossly impaired in vitro. Thus, the death of Brca1(5-6) mutant embryos prior to gastrulation may be due to a failure of the proliferative burst required for the development of the different germ layers.

Alternative Splicing↗

PubMed1995

Limb deformity proteins during avian neurulation and sense organ development.

The nuclear Limb deformity (Ld) proteins (formins) are expressed during the avian primitive streak stages. Initially, they are detected predominantly in cells of the forming notochord, scattered mesodermal precursors and the induced neural plate. No expression is detected in endodermal cells. The subsequent graded distribution of Ld positive cells along the anterior-posterior axis of the neural tube follows the antero-posterior progression of its differentiation. The Ld proteins are also differentially expressed during induction and development of both the inner ear and eye. An unequal distribution of Ld proteins along the dorso-ventral axis of the otic vesicle is observed during its initial patterning. In the eye, the Ld proteins are expressed by the optic vesicle during secondary induction of the lens placode. Following induction, the proteins are also expressed by the newly formed lens placode, a process which is reminiscent of homeogenetic induction. During differentiation of the retina and lens, the Ld domains seem to demarcate territories, giving rise to specific eye structures. A comparative analysis of the Ld distribution and BrdU incorporation in the two sense organs indicates that the proteins are predominantly expressed by committed and/or differentiating (post-mitotic) cells. In general, expression of Ld proteins is induced during determination and remains during differentiation of particular cell-types. This study implies that the nuclear Ld proteins are involved in morphogenesis of both neuro-ectodermal and mesodermal structures.

Animals↗