Pancreatic Homeodomain Transcription Factor IDX1/IPF1 Expressed in Developing Brain Regulates Somatostatin Gene Transcription in Embryonic Neural Cells

Abstract

Hox-like homeodomain proteins play a critical role during embryonic development by regulating the transcription of genes that are important for the generation of specific organs or cell types. The homeodomain transcription factor IDX1/IPF1, the expression of which was thought until recently to be restricted to the pancreas and foregut, is required for pancreas development and for the expression of genes controlling glucose homeostasis. We report that IDX1/IPF1 is also expressed in embryonic rat brain at a time coincident with active neurogenesis. Electrophoretic mobility shift assays with nuclear extracts of embryonic brains indicated that IDX1/IPF1 binds to two somatostatin promoter elements, SMS-UE-B and the recently discovered SMS-TAAT3. The requirement of these elements for IDX1/ IPF1 transactivation of the somatostatin gene in neural cells was confirmed in transfection studies using embryonic cerebral cortex-derived RC2.E10 cells. Immunohistochemical staining of rat embryos showed IDX1/IPF1-positive cells located near the ventricular surface in germinative areas of the developing central nervous system. Cellular colocalization of IDX1/IPF1 and somatostatin was found in several areas of the developing brain, including cortex, ganglionic eminence, hypothalamus, and inferior colliculus. These results support the notion that IDX1/IPF1 regulates gene expression during development of the central nervous system independent of its role on pancreas development and function.Homeodomain genes encode a large family of transcription factors that are grouped in different classes, including, for example, Hox-, POU-, Pax-, LIM-, and Dlx-like proteins, depending on the relative degree of conservation of the homeodomain. During gestation, expression of individual homeodomain genes is critical for proper regional development of the central nervous system (CNS) 1 and peripheral tissues of the embryo. IDX1/IPF1 (also known as STF-1 and PDX-1) is a transcription factor encoded by a Hox-like homeodomain gene, the expression of which was initially described to be restricted to the pancreas and duodenum, and it has been demonstrated that pancreas development requires IDX1/IPF1 in mice (1, 2). This requirement was confirmed in the human by the identification of an individual with pancreatic agenesis and a homozygous mutation in the IPF-1 gene (3). In addition, IDX1/IPF1 is essential for normal pancreatic islet function by regulating the expression of a number of pancreatic genes, including insulin, somatostatin, islet amyloid polypeptide, and glucose transporter type 2 (4 -8). Importantly, in humans, heterozygous mutations of the IPF-1 gene are linked to a type of autosomal dominant diabetes mellitus known as maturity onset diabetes of the young (MODY4) (9).In the developing neural tube, different homeodomain genes that control mechanisms of regional and cellular determination are selectively expressed in the primordia of the three major regions of the CNS: forebrain, midbrain, and hindbrain (10). In the hindbrain, certain Hox-like ...

The Homeoprotein Alx3 Contains Discrete Functional Domains and Exhibits Cell-specific and Selective Monomeric Binding and Transactivation

“…6B). Previously, we described that IDX1, a different homeodomain protein present in neural cells binds the SMS-UE-B element (35). Most interestingly, we found that the addition of two different anti-IDX1 antisera to the binding reaction resulted in the disappearance of the same band that disappears with the anti-Alx3 antiserum, suggesting that the protein complex bound to SMS-UE-B may contain both homeoproteins (Fig.…”

“…For comparison, we also used nuclear extracts of pancreatic isletderived somatostatin producing RIN-1027-B2 cells. We have previously shown that nuclear proteins from both RC2.E10 and RIN-1027-B2 cells bind to somatostatin TAAT-containing elements forming several sequence-specific protein-DNA complexes (18,33,35). Expression of Alx3 in RC2.E10 and RIN-1027-B2 cells was confirmed by Western blot (Fig 6A).…”

“…6B). This situation is similar to that reported for IDX1, which recognizes SMS-TAAT2 in somatostatin-producing pancreatic cells, but not in somatostatin-producing neural cells, despite the fact that it is expressed in both cell types (34,35). This prompted us to investigate whether Alx3 present in pancreatic cells binds SMS-TAAT1 or SMS-TAAT2.…”

“…Labeled products did not bind to GST or to GST-Alx3⌬N140C (not shown). E, binding of 35 S-labeled, in vitro translated Alx3-(143-228) to GST-Alx3-(143-228) and GST-Alx3 FL but not to GST or to GST-Alx3⌬N140C. In this case, the labeled Alx3-(143-228) polypeptide was resolved in a 20% polyacrylamide gel.…”

“…The promoter region of the somatostatin gene contains at least four homeodomain-binding DNA cis-regulatory elements with TAAT core motifs, known as SMS-UE-B, SMS-TAAT1, SMS-TAAT2, and SMS-TAAT3, the function of which has been studied in detail in pancreatic and neural cells (18,(33)(34)(35). In the case of the GFAP gene, studies on the regulation of its expression by homeodomain proteins have not been reported.…”