Аннотация
The nuclear matrix has been implicated in several cellular processes,
including DNA replication, transcription, and RNA processing. In
particular, transcriptional regulation is believed to be accomplished
by binding of chromatin loops to the nuclear matrix and by the concentration
of specific transcription factors near these matrix attachment regions
(MARs). A number of MAR-binding proteins have been identified, but
few have been directly linked to tissue-specific transcription. Recently,
we have identified two cellular protein complexes (NBP and UBP) that
bind to a region of the mouse mammary tumor virus (MMTV) long terminal
repeat (LTR) previously shown to contain at least two negative regulatory
elements (NREs) termed the promoter-proximal and promoter-distal
NREs. These NREs are absent from MMTV strains that cause T-cell lymphomas
instead of mammary carcinomas. We show here that NBP binds to a 22-bp
sequence containing an imperfect inverted repeat in the promoter-proximal
NRE. Previous data showed that a mutation (p924) within the inverted
repeat elevated basal transcription from the MMTV promoter and destabilized
the binding of NBP, but not UBP, to the proximal NRE. By using conventional
and affinity methods to purify NBP from rat thymic nuclear extracts,
we obtained a single major protein of 115 kDa that was identified
by protease digestion and partial sequencing analysis as the nuclear
matrix-binding protein special AT-rich sequence-binding protein 1
(SATB1). Antibody ablation, distamycin inhibition of binding, renaturation
and competition experiments, and tissue distribution data all confirmed
that the NBP complex contained SATB1. Similar types of experiments
were used to show that the UBP complex contained the homeodomain
protein Cux/CDP that binds the MAR of the intronic heavy-chain immunoglobulin
enhancer. By using the p924 mutation within the MMTV LTR upstream
of the chloramphenicol acetyltransferase gene, we generated two strains
of transgenic mice that had a dramatic elevation of reporter gene
expression in lymphoid tissues compared with reporter gene expression
in mice expressing wild-type LTR constructs. Thus, the 924 mutation
in the SATB1-binding site dramatically elevated MMTV transcription
in lymphoid tissues. These results and the ability of the proximal
NRE in the MMTV LTR to bind to the nuclear matrix clearly demonstrate
the role of MAR-binding proteins in tissue-specific gene regulation
and in MMTV-induced oncogenesis.
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