Affinity of linker histones for chromatin in situ analyzed using DAPI as a cytochemical probe.
BACKGROUND
It is generally assumed that linker histones contribute to condensation of chromatin and to the repression of genetic activity by binding to chromatin with increased affinity. The aim of this study is to investigate a possible correlation between linker histone binding to chromatin in situ and chromatin condensation using a cytochemical approach that largely preserves nuclear structure.


METHODS
Ionically bound H1 was extracted with increasing concentrations of sodium chloride. Thereafter, the cells were fixed in formaldehyde and stained with 50 nM 4',6-diamidino-2-phenylindole (DAPI) and fluorescence intensity was measured by image cytofluorometry.


RESULTS
The association between linker histones and chromatin was stronger in cultured human fibroblasts and rat smooth muscle cells than in both human T lymphocytes and granulocytes, and in particular, frog erythrocytes, which exhibited highly condensed chromatin. Our data indicate a lower affinity of linker histones for metaphase chromosomes than for interphase chromatin, and we observed a reduction in linker histone affinity during terminal differentiation of frog erythrocytes in vivo.


CONCLUSIONS
Taken together, our findings imply that the affinity of linker histones for chromatin in situ is unrelated or inversely related to chromatin condensation.
Loborg
H
H
Department of Biomedicine and Surgery, Division of Cell Biology, Faculty of Health Sciences, Linköping University, Linköping, Sweden. hello@mcb.liu.se
Rundquist
I
I
D000818:Animals; D001011:Aorta / Q000166:cytology; D002454:Cell Differentiation; D002455:Cell Division / Q000187:drug effects; D002467:Cell Nucleus / Q000737:chemistry / Q000378:metabolism; D002843:Chromatin / Q000737:chemistry / Q000378:metabolism*; D004912:Erythrocytes / Q000166:cytology; D005260:Female; D005347:Fibroblasts / Q000166:cytology; D016193:G1 Phase; D016195:G2 Phase; D006657:Histones / Q000032:analysis* / Q000378:metabolism*; D006801:Humans; D019044:Image Cytometry / Q000379:methods; D007211:Indoles; D007364:Intercalating Agents; D009131:Muscle, Smooth, Vascular / Q000166:cytology; D010659:Phenylhydrazines / Q000494:pharmacology; D011485:Protein Binding; D051381:Rats; D012965:Sodium Chloride / Q000494:pharmacology; D013045:Species Specificity; D013601:T-Lymphocytes / Q000166:cytology; D014982:Xenopus laevis
D002843:Chromatin; D006657:Histones; D007211:Indoles; D007364:Intercalating Agents; D010659:Phenylhydrazines; C030299:phenylhydrazine; D012965:Sodium Chloride; C007293:DAPI