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The octamer sequence ATGCAAAT is found in the promoters of immunoglobulin (Ig) heavy and light chain genes and in the heavy chain enhancer and is a major determinant of the cell type specific expression of Ig genes in B cells. An apparent paradox is that the same sequence serves as an upstream promoter or enhancer element in a variety of housekeeping genes such as the histone H2B and U snRNA genes. The differential usage of this regulatory sequence motif is thought to be mediated by different species of octamer binding proteins. One species of 100 kd, designated OTF‐1, is present in all cell types and may exert its activating function only when it can interact with additional adjacent transcription factors. The lymphoid cell specific octamer binding protein of 60 kd (OTF‐2A) specifically stimulates Ig promoters which consist essentially of a TATA‐box and an octamer sequence upstream of it. Here we present evidence for yet another B cell specific octamer binding protein of 75 kd (OTF‐2B). From several findings, including the absence of OTF‐2B (but not OTF‐2A) from a lymphocyte line that cannot respond to the IgH enhancer, we propose a role of the novel octamer factor in the long range activation by the IgH enhancer. We have used the proteolytic clipping bandshift assay (PCBA) technique to distinguish the three different forms found in B cells. This analysis indicates that the 75 kd‐species OTF‐2B is closely related to the 60 kd species OTF‐2A.

The 'octamer' sequence, ATGCAAAT or its complement ATTTGCAT, is a key element for the transcriptional regulation of immunoglobulin genes in B-lymphocytes as well as a number of housekeeping genes in all cell types. In lymphocytes, the octamer-binding protein Oct-2A and variants thereof are thought to contribute to the B-cell specific gene expression, while the ubiquitous protein Oct-1 seems to control general octamer site-dependent transcription. Various other genes, for example interleukin-1 and MHC class II genes, contain an octamer sequence in the promoter and are expressed in cells of both the immune and nervous systems. This prompted us to analyze the octamer-binding proteins in the latter cells. Using the electrophoretic mobility shift assay, at least six novel octamer binding proteins were detected in nuclear extracts of cultured mouse astrocytes. These proteins are differentially expressed in human glioblastoma and neuroblastoma cell lines. The nervous system-derived (N-Oct) proteins bound to the octamer DNA sequence in a manner which is indistinguishable from the Oct-1 and Oct-2A proteins. The relationship of the N-Oct proteins to Oct-1 and Oct-2A was analyzed by proteolytic clipping bandshift assays and by their reactivity towards antisera raised against recombinant Oct-1 and Oct-2A proteins. On the basis of these assays, all N-Oct-factors were found to be distinct from the ubiquitous Oct-1 and the lymphoid-specific Oct-2A proteins. In melanoma cells that contain the N-Oct-3 factor, a transfected lymphocyte-specific promoter was neither activated nor was it repressed upon contransfection with an Oct-2A expression vector. We therefore speculate that N-Oct-3 and other N-Oct factors have a specific role in gene expression in cells of the nervous system.

All promoters of immunoglobulin heavy chain genes contain three conserved sequence motifs: a heptamer motif CTCATGA, an octamer motif ATGCAAAT, and a TATA box. We show that, despite their different sequences, both the heptamer and the octamer motif are bound by the same octamer transcription factors (Oct factors, also referred to as OTFs), namely the lymphoid‐specific proteins Oct‐2A and Oct‐2B, as well as the ubiquitous protein Oct‐1. Even though binding to the octamer motif is stronger, a single heptamer motif can bind Oct proteins and mediate transcriptional activity in lymphoid cells. Furthermore, factor binding to the octamer motif facilitates binding to the nearby heptamer motif. We propose that the heptamer element plays a role early in B‐cell differentiation to ensure that the heavy chain promoters are transcriptionally activated before the light chain promoters, which do not contain the heptamer motif.

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