Immunolabelling of human metaphase chromosomes reveals the same banded distribution of histone H3 isoforms methylated at lysine 4 in primary lymphocytes and cultured cell lines.
BACKGROUND
Using metaphase spreads from human lymphoblastoid cell lines, we previously showed how immunofluorescence microscopy could define the distribution of histone modifications across metaphase chromosomes. We showed that different histone modifications gave consistent and clearly defined immunofluorescent banding patterns. However, it was not clear to what extent these higher level distributions were influenced by long-term growth in culture, or by the specific functional associations of individual histone modifications.


RESULTS
Metaphase chromosome spreads from human lymphocytes stimulated to grow in short-term culture, were immunostained with antibodies to histone H3 mono- or tri-methylated at lysine 4 (H3K4me1, H3K4me3). Chromosomes were identified on the basis of morphology and reverse DAPI (rDAPI) banding. Both antisera gave the same distinctive immunofluorescent staining pattern, with unstained heterochromatic regions and a banded distribution along the chromosome arms. Karyotypes were prepared, showing the reproducibility of banding between sister chromatids, homologue pairs and from one metaphase spread to another. At the light microscope level, we detect no difference between the banding patterns along chromosomes from primary lymphocytes and lymphoblastoid cell lines adapted to long-term growth in culture.


CONCLUSIONS
The distribution of H3K4me3 is the same across metaphase chromosomes from human primary lymphocytes and LCL, showing that higher level distribution is not altered by immortalization or long-term culture. The two modifications H3K4me1 (enriched in gene enhancer regions) and H3K4me3 (enriched in gene promoter regions) show the same distributions across human metaphase chromosomes, showing that functional differences do not necessarily cause modifications to differ in their higher-level distributions.
School of Cancer Sciences, College of Medical and Dental Sciences, University of Birmingham, Edgbaston, Birmingham, B15 2TT, UK. e.terrenoire@chu-tours.fr.|West Midlands Regional Genetics Laboratory, Birmingham Women's NHS Foundation Trust, Mindelsohn Way, Edgbaston, Birmingham, B15 2TG, UK. e.terrenoire@chu-tours.fr.|Present address : Service de Génétique, CHU de Tours, 2 Boulevard Tonnellé, 37044, Tours, Cedex 09, France. e.terrenoire@chu-tours.fr.
Edith
E
Terrenoire
School of Cancer Sciences, College of Medical and Dental Sciences, University of Birmingham, Edgbaston, Birmingham, B15 2TT, UK. j.halsall@bham.ac.uk.
John A
JA
Halsall
School of Cancer Sciences, College of Medical and Dental Sciences, University of Birmingham, Edgbaston, Birmingham, B15 2TT, UK. b.m.turner@bham.ac.uk.
Bryan M
BM
Turner
D002460:Cell Line; D002871:Chromosome Banding; D002877:Chromosomes, Human; D006657:Histones / Q000235:genetics* / Q000378:metabolism*; D006801:Humans; D008214:Lymphocytes / Q000378:metabolism*; D008239:Lysine; D008677:Metaphase; D008745:Methylation; D020033:Protein Isoforms
D006657:Histones; D020033:Protein Isoforms; D008239:Lysine