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J Nickerson

Publications and source records attributed to J Nickerson.

11 recordsLinked to original sources
PubMed1998

Intranuclear targeting of AML/CBFalpha regulatory factors to nuclear matrix-associated transcriptional domains.

The AML/CBFalpha runt transcription factors are key regulators of hematopoietic and bone tissue-specific gene expression. These factors contain a 31-amino acid nuclear matrix targeting signal that supports association with the nuclear matrix. We determined that the AML/CBFalpha factors must bind to the nuclear matrix to exert control of transcription. Fusing the nuclear matrix targeting signal to the GAL4 DNA binding domain transactivates a genomically integrated GAL4 responsive reporter gene. These data suggest that AML/CBFalpha must associate with the nuclear matrix to effect transcription. We used fluorescence labeling of epitope-tagged AML-1B (CBFA2) to show it colocalizes with a subset of hyperphosphorylated RNA polymerase II molecules concentrated in foci and linked to the nuclear matrix. This association of AML-1B with RNA polymerase II requires active transcription and a functional DNA binding domain. The nuclear matrix domains that contain AML-1B are distinct from SC35 RNA processing domains. Our results suggest two of the requirements for AML-dependent transcription initiation by RNA polymerase II are association of AML-1B with the nuclear matrix together with specific binding of AML to gene promoters.

Cell Compartmentation↗
PubMed1997

Cell cycle independent interaction of CDC2 with the centrosome, which is associated with the nuclear matrix-intermediate filament scaffold.

The cell cycle regulating Cdc2 protein kinase helps orchestrate cell cycle dependent changes in cell structure and function. This report shows that Cdc2 is localized to the centrosome region and is tightly bound to the nuclear matrix-intermediate filament scaffold. Antibodies to Cdc2 and to the centrosome-specific protein, pericentrin, label the centrosome in an apparently cell cycle independent manner. Isolated centrosomes also label similarly with both antibodies. Essentially, all cells show Cdc2 labeling of the centrosomes, implying an independence of the stage in the cell cycle, a conclusion supported by studies of synchronized cells. In contrast to the labeling of every cell with the Cdc2 monoclonal antibody, fewer centrosomes were labeled with an antibody to the PSTAIRE domain of Cdc2. Embedment-free, immunogold electron micrographs of extracted cell whole mounts show the centrioles and a pericentriolar network of filaments. Both Cdc2 and pericentrin antibodies decorate the amorphous pericentriolar material, while the Cdc2 antibodies also decorate the centrioles themselves. The constitutive presence of Cdc2 at the centrosome suggests a continuing role in the dynamics of centrosome function throughout the cell cycle.

CDC2 Protein Kinase↗
PubMed1995

The nuclear matrix protein NMP-1 is the transcription factor YY1.

NMP-1 was initially identified as a nuclear matrix-associated DNA-binding factor that exhibits sequence-specific recognition for the site IV regulatory element of a histone H4 gene. This distal promoter domain is a nuclear matrix interaction site. In the present study, we show that NMP-1 is the multifunctional transcription factor YY1. Gel-shift and Western blot analyses demonstrate that NMP-1 is immunoreactive with YY1 antibody. Furthermore, purified YY1 protein specifically recognizes site IV and reconstitutes the NMP-1 complex. Western blot and gel-shift analyses indicate that YY1 is present within the nuclear matrix. In situ immunofluorescence studies show that a significant fraction of YY1 is localized in the nuclear matrix, principally but not exclusively associated with residual nucleoli. Our results confirm that NMP-1/YY1 is a ubiquitous protein that is present in both human cells and in rat osteosarcoma ROS 17/2.8 cells. The finding that NMP-1 is identical to YY1 suggests that this transcriptional regulator may mediate gene-matrix interactions. Our results are consistent with the concept that the nuclear matrix may functionally compartmentalize the eukaryotic nucleus to support regulation of gene expression.

Animals↗
PubMed1994

An early decrease in interphotoreceptor retinoid-binding protein gene expression in Abyssinian cats homozygous for hereditary rod-cone degeneration.

Levels of interphotoreceptor retinoid-binding protein (IRBP) protein and message in retinas of Abyssinian cats homozygous for progressive rod-cone degeneration were determined at early ages, well before the onset of clinical retinal degeneration. IRBP gene expression was assessed by immunochemical quantitation of IRBP protein, and by Northern blotting and slot-blotting of total RNA using a human IRBP cDNA probe. Morphology was assessed by electron microscopy and immunocytochemistry. Levels of both IRBP protein and message in affected Abyssinian cat retinas were significantly reduced below normal as early as 4 weeks of age at the earliest stage of retinal disorientation. Opsin mRNA was more abundant in affected Abyssinian cat retinas than in control retinas. This was at least 1 year before the onset of clinical symptoms. The reduction in IRBP gene expression to levels significantly below normal well before the onset of retinal degeneration in affected Abyssinian cat retinas indicates that this represents a primary defect or at least an early problem that could itself cause adverse effects.

Animals↗
PubMed1993

Human retinoblastoma cells express alpha B-crystallin in vivo and in vitro.

alpha B-crystallin is a major lens protein that is a member of the heat-shock family of proteins. Using immunohistochemical and northern blot techniques, we now demonstrate its presence in freshly-fixed retinoblastoma tissue. The protein is also abundantly expressed in cultured human retinoblastoma cells (Y-79 NEI, WERI Rb-1) as well as two subcultured Y-79 lines (ATCC and GM01232C). High expression of alpha B-crystallin may be involved in tumor growth and/or be a marker for general oncogenic "stress" in the tumor tissue.

Animals↗
PubMed1992

Synthesis and secretion of interphotoreceptor retinoid-binding protein (IRBP) and developmental expression of IRBP mRNA in normal and rd mouse retinas.

The synthesis and secretion of interphotoreceptor retinoid-binding protein (IRBP) was quantitatively assessed in retinas of normal and rd mutant mice using short-term organ culture with [35S]methionine. Retinas were studied at ages P9-P12, time points prior to and immediately after the onset of the degeneration of the rd retina. Soluble proteins of the retinal pellet and the incubation medium were subjected to SDS-polyacrylamide gel electrophoresis. Analysis of labeled protein bands utilized a radioactivity scanning system to quantify [35S]methionine incorporation into newly synthesized IRBP. The synthesis and secretion into the incubation medium of IRBP by rd mouse retinas was comparable to normal retinas at P9-P10 but decreased by more than 50% by P12. IRBP mRNA levels were evaluated in retinas of normal and rd mice ages P7-P14. Although IRBP mRNA expression increased in the rd mouse through P10, it decreased markedly thereafter. Previously reported immunocytochemical studies suggested that IRBP was not secreted in the rd mouse retina. The results of this study indicate, however, that rd mouse retinas, when removed from the eye, have the capacity to synthesize and secrete IRBP.

Animals↗
PubMed1992

A molecular perspective on mammalian evolution from the gene encoding interphotoreceptor retinoid binding protein, with convincing evidence for bat monophyly.

The evolutionary relationships of the various orders of placental mammals remain an issue of uncertainty and controversy. Molecular studies of mammalian phylogeny at the DNA level that include more than just a few orders are still relatively meager. Here we report results on mammalian phylogeny deduced from the coding sequence of the single-copy nuclear gene for the interphotoreceptor retinoid binding protein (IRBP). Analysis of 13 species representing eight eutherian orders and one marsupial yielded results that falsify the hypothesis that megachiropteran bats are "flying primates," only convergently resembling microchiropteran bats. Instead, in agreement with more traditional views, as well as those from other recent molecular studies, the results strongly support a monophyletic Chiroptera (micro- and megabats grouped together). The IRBP results also offer some rare molecular support for the Glires concept, in which rodents and lagomorphs form a superordinal grouping. Also in congruence with other recent molecular evidence, IRBP sequences do not support the view of a superorder Archonta that includes Chiroptera along with Dermoptera (flying lemur), Scandentia (tree shrew), and Primates. IRBP was not however, without its shortcomings as a molecular phylogenetic system: high levels of homoplasy, evident in the marsupial outgroup, did not allow us to properly root the tree, and several of the higher level eutherian clades were only weakly supported (e.g., a Carnivora/Chiroptera clade and an Artiodactyla/Carnivora/Chiroptera clade). We suggest that these shortcomings may be diminished as the phylogenetic density of the data set is increased.

Amino Acid Sequence↗
PubMed1991

Human papillomavirus 16 E7 protein is associated with the nuclear matrix.

The cellular localization of the human papillomavirus (HPV)-16 E7 gene product in the cell lines CaSki and SiHa has been determined by both biochemical and immunocytochemical methods. These measurements show E7 to be localized in the cell nucleus, specifically with the nonchromatin nuclear structure or nuclear matrix. This localization of E7 required an unambiguous fractionation of the nuclear constituents. This was achieved by using a gentle sequential fractionation procedure to prepare the scaffold consisting of the nuclear matrix and intermediate filaments (NM-IF). Chromatin was cleaved with nuclease and the resulting nucleosomes eluted with 0.25 M ammonium sulfate. Immunostaining of cells after this extraction procedure with monoclonal antibodies (mAbs) to E7 revealed a fine grained, punctate nuclear fluorescence in CaSki and SiHa, which was absent in normal cervical keratinocytes and the HPV-negative cell line C33.1. Western blots of cell fractions with these mAbs showed that E7 was localized in the NM-IF fraction in SiHa and CaSki but was not detected in HPV-negative cells. A second protein of slightly higher mobility is identified by these antisera in HPV-16-containing cells. The data suggest that the previous inability to directly visualize E7 by immunocytology is due to the masking of epitopes by cellular components and not to low levels of protein.

Blotting, Western↗
PubMed1990

Localization of the gene for interphotoreceptor retinoid-binding protein to mouse chromosome 14 near Np-1.

Interphotoreceptor retinoid-binding protein (IRBP) is a large glycoprotein known to bind retinoids and found primarily in the interphotoreceptor matrix of the retina between the retinal pigment epithelium and the photoreceptor cells. It is thought to transport retinoids between the retinal pigment epithelium and the photoreceptors, a critical role in the visual process. We have used a 900-bp bovine IRBP cDNA fragment to map the corresponding gene, Rbp-3, to mouse chromosome 14 with somatic cell hybrids and have positioned the gene near Np-1 (nucleoside phosphorylase-1) by analysis of the progeny of an intersubspecific backcross. In the human genome, NP maps to human chromosome 14 and RBP3 to human chromosome 10. Thus, these two genes span the putative site of a chromosomal translocation which contributed to divergent karyotype evolution of man and mouse.

Animals↗