Identification of SP3 as a Negative Regulatory Transcription Factor in the Monocyte Expression of Growth Hormone*

Abstract

A number of studies from different laboratories clearly show that cells of the immune system produce a GH molecule indistinguishable from that produced in the pituitary. A more recent finding from our studies suggests that monocytes use the same first exon and promoter sequence for the expression of lymphocyte GH as that reported for the expression of pituitary GH. In this report we have extended these results by determining that two members of the SP family of transcription factors, SP1 and SP3, bind to the region at -138/-133 bp containing a GGGAGG motif. Confirmation that this region of the monocyte GH promoter-bound SP1 and SP3 was accomplished using electrophoretic mobility shift assays with SP1 consensus and mutant probes as well as specific antibodies to SP1 and SP3. Selective mutation of the SP1/SP3 site increased basal transcription by 73%, indicating that this site is important in transcriptional inhibition. Overexpression of SP1 had no demonstrable effect on the GH promoter, whereas overexpression of SP3 caused inhibition of expression in P-388 monocyte cells. Cotransfection of P-388 cells with overexpression vectors for both SP1 and SP3 transcription factors also resulted in inhibition of basal expression. Transfection experiments in Drosophila SL-2 cells overexpressing SP1 and/or SP3 suggest that both factors repress the basal expression of GH promoter luciferase constructs and that the effect together was additive. Taken together, the results demonstrate that basal expression of monocyte GH may be negatively regulated by SP3.

A homozygous point mutation in the GH1 promoter (c.-223C>T) leads to reduced GH1 expression in siblings with isolated GH deficiency (IGHD)

“…The transcriptional factor SP1 may act as either activator or repressor of gene expression depending on numerous variants as cell type, concentration of other intracellular proteins such as SP3 and post-translational modifications of SP1 protein, i.e. glycosylation, phosphorylation and sumoylation (8,22). Although SP1 is considered a repressor of GH1 expression, as interaction between SP1 and its binding site located at c.-163 of GH1 promoter decreases GH1 expression (8), interaction between SP1 and the binding site located at c.-223 was not previously studied, and at this position, SP1 could act stimulating GH1 expression.…”

“…glycosylation, phosphorylation and sumoylation (8,22). Although SP1 is considered a repressor of GH1 expression, as interaction between SP1 and its binding site located at c.-163 of GH1 promoter decreases GH1 expression (8), interaction between SP1 and the binding site located at c.-223 was not previously studied, and at this position, SP1 could act stimulating GH1 expression. Moreover, in silico analysis may have not identified other activator transcriptional factors that bind to GH1 promoter at c.-223.…”

“…The promoter region is highly polymorphic, and at least 16 single nucleotide polymorphisms (SNPs) have been reported within a 535-basepair segment upstream of the transcription start site (5,6,7). The interaction between the promoter region and activating transcription factors (such as POU1F1 and NF1) or repressive factors (such as SP1) leads to the activation or repression of GH1 expression respectively (8,9,10,11).…”

Age-Associated Loss of Bone Marrow Hematopoietic Cells Is Reversed by GH and Accompanies Thymic Reconstitution

“…For example, GH is synthesized by a variety of lymphoid organs (45,46), and this locally produced GH may act directly on these tissues (47). Endocrine and paracrine GH may also increase IGF-I that is produced by cells in both the thymus (48) and bone marrow (34).…”

Specificity protein-1 and -3 trans-activate the ovine placental lactogen gene promoter

“…Various reports have shown that Sp1 and Sp3 can act either synergistically to stimulate transcription, or may in fact compete for binding when both are present (Bouwman et al, 2000; Wong and Lee, 2002; Yu et al, 2003). There is evidence that Sp3 may repress Sp1 mediated transactivation, depending on the ratio of Sp1 to Sp3, the number of binding sites for Sp proteins within the promoter, or the tissue in which they are expressed (Vines and Weigent, 2000; Yu et al, 2003). While both Sp1 and Sp3 were shown to interact with FP6 in electrophoretic mobility supershift analysis of BeWo and oBNC nuclear extracts, and the use of over-expression and RNA interference constructs demonstrated functional interaction of both Sp1 and Sp3 with FP6 in BeWo cells, Western and Southwestern analysis of oBNC nuclear extracts suggested that Sp3 is the predominant factor in the sheep placenta.…”