Comparison of DNA-protein interactions in intact nuclei from avian liver and erythrocytes: a cross-linking study.
DNA-protein cross-linkages were formed in intact nuclei of chicken erythrocytes and liver cells by the action of cis-diammine dichloroplatinum (II). Most cross-linked proteins were components of the nuclear matrix, and their heterogeneity reflected the different complexity of liver and erythrocytes matrices, respectively. Some basic proteins, including histones, were also cross-linked, particularly in erythrocyte nuclei. South-Western blotting revealed that a variety of proteins isolated from the cross-linked liver nuclei recognized DNA specifically. In this group of proteins two relatively abundant, acidic, species of 38 and 66 kDa, respectively, might represent novel DNA-binding proteins from the nuclear matrix. In the case of erythrocytes, only the basic proteins showed a DNA-recognition capacity, and among them there were some unidentified species, absent from liver. Lamin B2 was cross-linked but was unable to recognize DNA, and the same was true for other abundant, cross-linked proteins from both types of nuclei. This led to the hypothesis that for some DNA-nuclear matrix interactions the aggregation typical of matrix proteins is essential for the specificity of DNA recognition. Hybridization analysis of the DNA isolated from the cross-linked complexes showed that SARs (scaffold attachment regions) and telomeric sequences were well represented in the cross-linked fragments, that the cross-linked DNA of liver was partially different from that of erythrocytes and that two defined SAR sequences were found to be present only in the cross-linked DNA. These results are in agreement with the present views on DNA-nuclear matrix interactions, which are usually studied on isolated nuclear matrices or purified proteins. Instead, our results provide experimental evidence obtained directly from intact nuclei.
Ferraro
A
A
Department of Biochemical Sciences, University La Sapienza, Rome, Italy.
Cervoni
L
L
Eufemi
M
M
Altieri
F
F
Turano
C
C
D000818:Animals; D002467:Cell Nucleus / Q000378:metabolism; D002645:Chickens; D015342:DNA Probes / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D015180:Electrophoresis, Gel, Two-Dimensional; D004912:Erythrocytes / Q000166:cytology* / Q000378:metabolism; D008099:Liver / Q000166:cytology* / Q000378:metabolism; D015530:Nuclear Matrix / Q000378:metabolism*
D015342:DNA Probes; D004268:DNA-Binding Proteins