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N Heintz

Publications and source records attributed to N Heintz.

At least 55 records · Page 3Linked to original sources
PubMed1994

Brain lipid-binding protein (BLBP): a novel signaling system in the developing mammalian CNS.

Using a polyclonal antibody against postnatal cerebellar cells, we have isolated a new, brain-specific member of the lipid-binding protein family (BLBP). Members of this family, such as cellular retinoic acid-binding protein, have been shown to carry small hydrophobic signaling molecules between cellular compartments. The expression of BLBP is spatially and temporally correlated with neuronal differentiation in many parts of the mouse CNS, including postnatal cerebellum, embryonic spinal cord, and cerebral cortex. In situ hybridization and immunocytochemistry show that BLBP is transiently expressed in radial glia in both the embryonic ventricular zone and the postnatal cerebellum. Subcellular localization studies by immunoelectron microscopy demonstrate that BLBP is present in the nucleus as well as the cytoplasm. Affinity-purified anti-BLBP antibodies block glial and neuronal differentiation in primary cell cultures, but have no effect on cell proliferation or adhesion. Based on these results, we propose that BLBP is required for the establishment of the radial glial fiber system in developing brain, a system that is necessary for the migration of immature neurons to establish cortical layers.

Amino Acid Sequence↗
PubMed1994

H1TF2A, the large subunit of a heterodimeric, glutamine-rich CCAAT-binding transcription factor involved in histone H1 cell cycle regulation.

H1TF2 is a CCAAT transcription factor that binds to the histone H1 subtype-specific consensus sequence, which has previously been shown to be necessary for temporal regulation of histone H1 transcription during the cell cycle (F. La Bella, P. Gallinari, J. McKinney, and N. Heintz, Genes Dev. 3:1982-1990, 1989). In this study, we report that H1TF2 is a heteromeric CCAAT-binding protein composed of two polypeptide doublets of 33 and 34 kDa and 43 and 44 kDa that are not antigenically related. The 33- and 34-kDa species were not detected in our previous studies (P. Gallinari, F. La Bella, and N. Heintz, Mol. Cell. Biol. 9:1566-1575, 1989) because of technical problems in detection of these heavily glycosylated subunits. The cloning of H1TF2A, the large subunit of this factor, reveals it to be a glutamine-rich protein with extremely limited similarity to previously cloned CCAAT-binding proteins. A monospecific antiserum produced against bacterially synthesized H1TF2A was used to establish that HeLa cell H1TF2A is phosphorylated in vivo and that, in contrast to the H2b transcription factor Oct1 (S. B. Roberts, N. Segil, and N. Heintz, Science 253:1022-1026, 1991; N. Segil, S. B. Roberts, and N. Heintz, Cold Spring Harbor Symp. Quant. Biol. 56:285-292, 1991), no gross change in H1TF2A phosphorylation is evident during the cell cycle. Further immunoprecipitation studies demonstrated that H1TF2 is heterodimeric in the absence of DNA in vivo and identified several H1TF2-interacting proteins that may play a role in H1TF2 function in vivo.

Amino Acid Sequence↗
PubMed1994

Kv3.3b: a novel Shaw type potassium channel expressed in terminally differentiated cerebellar Purkinje cells and deep cerebellar nuclei.

A two-step hybridization/subtraction procedure was employed to isolate markers for the later stages of Purkinje cell differentiation. From this screen, a novel Shaw potassium channel cDNA (Kv3.3b) was identified that is developmentally regulated. Expression of this channel is highly enriched in the brain, particularly in the cerebellum, where its expression is confined to Purkinje cells and deep cerebellar nuclei. Sequence analysis revealed that it is an alternatively spliced form of the mouse Kv3.3 gene, and that the previously reported Kv3.3 mRNA (Ghanshani et al., 1992) is not expressed in cerebellum. Expression of the Kv3.3b mRNA begins in cerebellar Purkinje cells between postnatal day 8 (P8) and P10 and continues through adulthood, coinciding with elaboration of the mature Purkinje cell dendritic arbor. The timing of expression of Kv3.3b mRNA is maintained in mixed, dissociated primary cerebellar cell culture. These results suggest that the Kv3.3b K+ channel function is restricted to terminally differentiated Purkinje cells, and that analysis of the mechanisms governing its expression in vivo and in vitro can reveal molecular mechanisms governing Purkinje cell differentiation.

Amino Acid Sequence↗
PubMed1993

RIP60 dimers and multiples of dimers assemble link structures at an origin of bidirectional replication in the dihydrofolate reductase amplicon of Chinese hamster ovary cells.

We show assembly of low and high multimers of HeLa cell nuclear protein, RIP60, at the origin of bidirectional replication (OBR) identified by Burhans, Vassilev, Caddle, Heintz and DePamphilis in Chinese hamster ovary cells. RIP60 binds a 5'-ATT-3' reiterated sequence downstream of the OBR and a second, homologous ATT sequence of opposite orientation situated within the OBR zone. Specifically bound structures were studied by conventional electron microscopy (EM) and quantitative scanning transmission electron microscopy (STEM). Dimers and multiples of dimers link the downstream binding site that overlaps a bent DNA sequence and the homologous upstream OBR sequence, looping out 700 bp of intervening DNA. Superposed dimers are found at individual unlinked sites, stabilized presumably through protein-protein interaction, and such superposition appears to occur also in the basic link structure. Along the loop, single crossovers and extended twists are observed by conventional EM. By STEM, loop DNA is laterally compacted, with diameter and mass equivalent to double-duplex DNA strands. Supercoiled 736 bp and 5243 bp circular DNAs assume similar laterally compacted geometries that are mostly absent from relaxed forms. These observations parallel the compacted, interwound superhelices viewed by cryo-electron microscopy in vitrified solutions containing magnesium ions, and provide structural evidence in agreement with that from conventional EM for superhelical tension in RIP60 loop DNA. Loop superhelicity could arise as a topological response to linking and suggests a functional role for link formation.

Animals↗
PubMed1993

Negative regulation of FAR1 at the Start of the yeast cell cycle.

In budding yeast, a switch between the mutually exclusive pathways of cell cycle progression and conjugation is controlled at Start in late G1 phase. Mating pheromones promote conjugation by arresting cells in G1 phase before Start. Pheromone-induced cell cycle arrest requires a functional FAR1 gene. We have found that FAR1 transcription and protein accumulation are regulated independently during the cell cycle. FAR1 RNA and protein are highly expressed in early G1, but decline sharply at Start. Far1 is phosphorylated just before it disappears at Start, suggesting that modification may target Far1 for degradation. Although FAR1 mRNA levels rise again during late S or G2 phase, reaccumulation of Far1 protein to functional levels is restricted until after nuclear division.

Base Sequence↗
PubMed1993

Changing patterns of gene expression define four stages of cerebellar granule neuron differentiation.

Among CNS neuronal populations, the cerebellar granule cell provides a simple model for analysing the molecular regulation of CNS neurogenesis. In this study, polyclonal antisera raised against immature granule cell precursors, purified from early postnatal mouse cerebellum, were used to isolate 39 unique cDNA clones from a lambda gt11 cDNA expression library made from the same cell population. Northern blot analysis revealed developmental stage and tissue-specific expression of 28 of the clones. In situ localization of mRNAs encoded by these novel cDNAs, as well as those encoding the axonal glycoprotein TAG-1 and the alpha 6 subunit of the GABAA receptor, reveal four distinct stages in cerebellar granule cell differentiation. The developmentally transient and spatially restricted expression of clones GC9 and GC44 identify a previously unrecognized step in cerebellar histogenesis.

Animals↗
PubMed1992

Squamous cell carcinoma of the cervix: HPV 16 and DNA ploidy as predictors of survival.

In this study, the hypothesis that DNA ploidy and the presence of HPV 16 and HPV 18 DNA affects survival of patients with squamous cell carcinoma of the cervix was tested. Archival paraffin blocks from biopsy and surgical specimens were obtained from 127 women diagnosed in 1977-1984. Determination of DNA ploidy was by flow cytometry and HPV 16 and HPV 18 DNA status by polymerase chain reaction with subsequent dot-blot hybridization. For each patient, age, stage, treatment modality, and 5-year survival were correlated with ploidy and HPV status. HPV 16 DNA was present in 53% of the tumors. HPV 18 was not detected in this population. HPV 16 DNA was found twice as often in Stages IB and IIA than in advanced-stage disease (III and IV). These advanced-stage tumors were more commonly aneuploid. Neither HPV status nor DNA ploidy were predictive of survival for any stage of disease or therapeutic modality.

Adult↗
PubMed1992

Eukaryotic replication origins as promoters of bidirectional DNA synthesis.

Recent work in yeast shows that eukaryotic origins of DNA replication are multipartite regulatory elements resembling promoters of transcription. As for the regulation of transcription, accessory transcription factors appear to function in concert with basic origin recognition factors to regulate initiation of DNA synthesis at specific subsets of origins. The participation of transcription factors in the regulation of DNA replication may facilitate temporal control of transcription and replication during the cell cycle, as well as providing a mechanism for integrating origin selection with the cellular transcriptional program.

DNA Mutational Analysis↗
PubMed1991

Mitotic phosphorylation of the Oct-1 homeodomain and regulation of Oct-1 DNA binding activity.

Oct-1 is a transcription factor involved in the cell cycle regulation of histone H2B gene transcription and in the transcription of other cellular housekeeping genes. Oct-1 is hyperphosphorylated as cells enter mitosis, and mitosis-specific phosphorylation is reversed as cells exit mitosis. A mitosis-specific phosphorylation site in the homeodomain of Oct-1 was phosphorylated in vitro by protein kinase A. Phosphorylation of this site correlated with inhibition of Oct-1 DNA binding activity in vivo and in vitro. The inhibition of Oct-1 DNA binding during mitosis suggests a mechanism by which the general inhibition of transcription during mitosis might occur.

Amino Acid Sequence↗
PubMed1991

Differential phosphorylation of the transcription factor Oct1 during the cell cycle.

Orderly progression through the somatic cell division cycle is accompanied by phase-specific transcription of a variety of different genes. During S phase, transcription of mammalian histone H2B genes requires a specific promoter element and its cognate transcription factor Oct1 (OTF1). A possible mechanism for regulating histone H2B transcription during the cell cycle is direct modulation of Oct1 activity by phase-specific posttranslational modifications. Analysis of Oct1 during progression through the cell cycle revealed a complex temporal program of phosphorylation. A p34cdc2-related protein kinase that is active during mitosis may be responsible for one mitotic phosphorylation of Oct1. However, the temporally controlled appearance of Oct1 phosphopeptides suggests the involvement of multiple kinases and phosphatases. These results support the idea that cell cycle-regulated transcription factors may be direct substrates for phase-specific regulatory enzymes.

CDC2 Protein Kinase↗
PubMed1991

Characterization and functional expression of a cDNA encoding egasyn (esterase-22): the endoplasmic reticulum-targeting protein of beta-glucuronidase.

Egasyn (esterase-22), a member of the nonspecific carboxylesterase multigene family (E.C. 3.1.1.1), is the endoplasmic reticulum (ER)-targeting protein of beta-glucuronidase. We utilized the polymerase chain reaction (PCR) in the eventual isolation of murine egasyn cDNAs. PCR primers were based upon: (1) partial amino acid sequences derived from egasyn peptides and (2) a conserved active site region shared by carboxylesterases. The amino acid sequence deduced from the PCR product matched that obtained from egasyn protein. This product was utilized as a probe to screen a cDNA library. Two cDNAs whose composite sequence encoded an open reading frame of 562 amino acids were isolated. A message size of 1700-2000 bp was revealed by RNA blot hybridization analysis. S1 nuclease protection analyses detected mRNA in liver, kidney, lung, and submandibular gland, but not in spleen, brain, and testes. Genetic mapping confirmed the location of an egasyn cDNA fragment in cluster 1 of the esterase region on chromosome 8. Transfection of COS cells with the 2022-bp cDNA resulted in the expression of esterase activity, which comigrated on native gels with liver esterase-22. The features of the deduced amino acid sequence of the egasyn cDNA are compared with previously characterized carboxylesterases and with other lumenal ER proteins.

Amino Acid Sequence↗
PubMed1991

Genetic mapping of the lurcher locus on mouse chromosome 6 using an intersubspecific backcross.

The lurcher (Lc) mutant mouse strain exhibits postnatal degeneration of cerebellar Purkinje cells. We have typed progeny from an intersubspecific, phenotypic backcross at seven loci to develop a genetic linkage map which spans approximately 35 cM surrounding and including the Lc locus on mouse chromosome 6. [(Mus musculus castaneus x B6CBA-Aw-J/A-Lc)F1 x B6CBA-Aw-J/A]N2 progeny were scored visually for the lurcher phenotype and molecularly, through restriction fragment length polymorphism analysis, for six cloned markers. Two candidate genes, Npy and Pcp-1, which map to mouse chromosome 6 and which are expressed in the cerebellum, are demonstrated to be distinct from Lc. Three genes are shown to be closely linked to the Lc locus, and the map order cen-Cpa-Npy-Cbl-1-Lc-Igk, Fabpl-Pcp-1 is determined. The molecular genetic linkage map presented here represents progress toward isolating a clone of the Lc gene.

Animals↗
PubMed1991

Genetic mapping of meander tail, a mouse mutation affecting cerebellar development.

The meander tail mouse harbors a recessive mutation on chromosome 4 that affects the anterior lobes of the cerebellum and the caudal vertebrae. Examination of the mea/mea cerebellum reveals that the complete disorganization of all cell types seen in the anterior lobes is separated by a sharp and consistent boundary from the normal cytoarchitecture of the posterior lobes. In the absence of any biochemical information regarding the affected gene product, attempts to clone the gene must rely on the strategy of reverse genetics. As an initial step in this process we have constructed a genetic linkage map spanning 68 cM of chromosome 4 using an intersubspecific phenotypic backcross. The loci included in this analysis are Calb, Ggtb, Lv, b, Ifa, mea, D4Rp1, Glut-1, Lck, Lmyc-1, and Eno-1. This analysis positions the mea phenotypic locus in the interval between Ifa and Glut1. These results also further define regions of homology between mouse chromosome 4 and human chromosomes 8, 1, and 9. This linkage map provides the means to evaluate candidate genes, and to identify tightly linked markers useful for cloning the meander tail locus.

Animals↗
PubMed1991

A multilocus linkage map of mouse chromosome 8.

We present a genetic linkage map of mouse chromosome 8 that spans 53 cM and includes eight cloned loci. This map was derived from analysis of 100 progeny of an interspecific backcross between Mus spretus and Mus musculus domesticus. Genes that were mapped in this analysis include L7, Plat, Lpl, Ucp, Es, Mt-1, Um, and Tat. This analysis positions a new extremely proximal marker on chromosome 8, which is discussed as a potential candidate gene for the nervous locus. These linkage data will be useful for the mapping of additional loci on chromosome 8.

Animals↗
PubMed1991

Cerebellar granule cell neurogenesis is regulated by cell-cell interactions in vitro.

When CNS precursor cells purified from the external germinal layer of the early postnatal mouse cerebellum are cultured in cellular reaggregates, DNA synthesis increased 10-fold above that of cells dispersed in a monolayer or embedded in a collagen matrix. Dividing precursor cells gave rise to neurons immunopositive for the neural antigens N-CAM, L1, and TAG-1, but not to astroglial cells immunopositive for glial filament protein. Moreover, proliferating precursor cells did not generate other types of cerebellar neurons, as judged by the lack of expression of glutamic acid decarboxylase, the synthetic enzyme for gamma-amino-n-butyric acid. By contrast, the addition of astroglial cells, or astroglial cell membranes, to cellular reaggregates of granule cell neuroblasts arrested precursor cell DNA synthesis in a dose-dependent manner. These results suggest that homotypic contact interactions among CNS neural progenitors control precursor cell proliferation and fate in generative zones of developing brain.

Aminobutyrates↗
PubMed1991

Transcriptional regulation in the eukaryotic cell cycle.

Cell-cycle-regulated transcription is a characteristic feature of complex cell cycles in organisms as divergent as yeasts and humans. Increasing evidence suggests that transcriptional regulation may control key events in the eukaryotic cell cycle. In this review we will address the mechanisms that may regulate transcription during the cell cycle and the roles which periodic transcription may serve in the control of cell cycle progression.

Cell Cycle↗