Regulation of rat haptoglobin gene expression is coordinated by the nuclear matrix.
Using computer stress-induced duplex destabilization (SIDD) analysis and binding experiments, we identified a S/MAR element (-599/-200 bp) (Hp-S/MAR) adjacent to the cis-element (-165/-56 bp) in the rat haptoglobin gene. We examined its functional interactions with the lamins and lamin-associated proteins in the basal state and during acute-phase (AP) response-induced increased transcription. Colocalization, electrophoretic mobility shift assay (EMSA), and re-electrophoresis of nucleoprotein complexes, South-Western and Western blot analysis and coimmunoprecipitation experiments revealed that the lamins, PARP-1, C/EBP beta, and Hp-S/MAR assembled higher order complexes through direct lamin-Hp-S/MAR and probably PARP-1-Hp-S/MAR interactions although C/EBP beta did not bind to the Hp-S/MAR but established direct interaction with PARP-1. The transition from constitutive to increased haptoglobin gene transcription during the AP response was associated with quantitative and qualitative changes in Hp-S/MAR-protein interactions, respectively, observed as increased association of the lamin(s) with the Hp-S/MAR and as the appearance of a 90 kDa Hp-S/MAR-binding protein. Also, during the AP response the contact between C/EBP beta and PARP-1 established in the basal state was lost. DNA chromatography with the haptoglobin cis-element and Western blot analysis suggests that PARP-1 was a coactivator during constitutive and elevated transcription. The results show that the lamin components of the nuclear matrix form a network of functional, dynamic protein-protein and protein-Hp-S/MAR associations with multiple partners, and underline the involvement of PARP-1 in the regulation of haptoglobin gene transcription. We concluded that the interplay of these interactions fine tunes haptoglobin gene expression to meet the changing requirements of liver cells.
Vidaković
Melita
M
Department of Molecular Biology, Institute for Biological Research, University of Belgrade, Bulevar Despota Stefana 142, 11060 Belgrade, Serbia. melita@ibiss.bg.ac.yu
Dinić
Svetlana
S
Grdović
Nevena
N
Mihailović
Mirjana
M
Uskoković
Aleksandra
A
Quesada
Piera
P
Poznanović
Goran
G
D000818:Animals; D022782:CCAAT-Enhancer-Binding Protein-beta / Q000378:metabolism; D005786:Gene Expression Regulation; D006242:Haptoglobins / Q000235:genetics*; D034882:Lamins / Q000378:metabolism; D008099:Liver / Q000166:cytology / Q000378:metabolism; D045170:Matrix Attachment Regions; D015530:Nuclear Matrix / Q000502:physiology*; D000071137:Poly (ADP-Ribose) Polymerase-1; D011065:Poly(ADP-ribose) Polymerases / Q000502:physiology*; D051381:Rats; D014158:Transcription, Genetic
D022782:CCAAT-Enhancer-Binding Protein-beta; D006242:Haptoglobins; D034882:Lamins; C499322:Parp1 protein, rat; D000071137:Poly (ADP-Ribose) Polymerase-1; D011065:Poly(ADP-ribose) Polymerases