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.
Department of Biochemical Sciences, University La Sapienza, Rome, Italy.
A
A
Ferraro
L
L
Cervoni
M
M
Eufemi
F
F
Altieri
C
C
Turano
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