Distinct factors with Sp1 and NF-A specificities bind to adjacent functional elements of the human U2 snRNA gene enhancer.

Abstract

The enhancer regions of mammalian and avian U1 and U2 small nuclear RNA (snRNA) genes are unusual in containing the sequence GGGCGG (GC-box) immediately upstream from the sequence ATGCAAAT (octamer). We made point mutations in the human U2 snRNA enhancer and tested them for the ability to direct U2 transcription in HeLa cells, as well as for the ability to form complexes with factors present in HeLa cell nuclear extracts. We show that neither the GC-box nor the octamer alone is sufficient for enhancer activity in vivo. Mutations in the GC-box reduce the ability of enhancer DNA fragments to bind a factor (probably Spl), whereas mutations in the octamer independently reduce the ability to bind a second factor (probably nuclear factor A, NF-A). The results suggest that adjacent binding of Spl and NF-A is an important feature of some UsnRNA gene enhancers.[Key Words: U2 snRNA~ U2 enhancer~ enhancer binding proteins; transcription factors]

Broad binding‐site specificity and affinity properties of octamer 1 and brain octamer‐binding proteins

“…The ubiquitously expressed protein octamer 1 (Oct-1) is a transcription factor having remarkably flexible DNA-sequence recognition properties. Oct-1 has been found to be involved in stimulating RNA-polymerase-I1 transcription of cellular genes that are constitutively active (Mattaj et al, 1985;Ares et al, 1987), cell-cycle regulated (LaBella et al, 1988), tissue-specific (LeBowitz et al, 1988;Johnson et al, 1990;Kemler et al, 1991) or developmentally regulated (Zwartkruis et al, 1992). It has also been shown to be involved in RNA-polymerase- Oct, octamer; POU, Pit-l-Oct-l-Unc-l ; ds, double-stranded.…”

Both Oct-1 and Oct-2A contain domains which can activate the ubiquitously expressed U2 snRNA genes.

“…The basic enhancer unit of the human U2 snRNA gene contains Spl binding sites combined with an octamer sequence (Janson et al, 1987(Janson et al, , 1989Ares et al, 1987). Transient expression experiments in HeLa cells have shown that there is an absolute requirement of the octamer motif for U2 transcription (Ares et al, 1987;Janson et al, 1989). Although an Spl binding site alone is unable to stimulate transcription, its deletion results in a 5-fold reduction in transcription (Janson et al, 1989).…”

“…Transcription of human U2 genes is controlled by a proximal sequence element (PSE) and a distal sequence element (DSE) which is also called the U2 enhancer (Westin et al, 1984;Ares et al, 1985;Mangin et al, 1986; for review see Dahlberg and Lund, 1987). The basic enhancer unit of the human U2 snRNA gene contains Spl binding sites combined with an octamer sequence (Janson et al, 1987(Janson et al, , 1989Ares et al, 1987). Transient expression experiments in HeLa cells have shown that there is an absolute requirement of the octamer motif for U2 transcription (Ares et al, 1987;Janson et al, 1989).…”

“…The octamer site is also important for transcription of the snRNA genes although their expression is not restricted to lymphoid cells (Mattaj et al, 1985;Ares et al, 1987;Ciliberto et al, 1987;Janson et al, 1987Janson et al, , 1989Dahlberg and Schenborn, 1988;Tanaka et al, 1988). Transcription of human U2 genes is controlled by a proximal sequence element (PSE) and a distal sequence element (DSE) which is also called the U2 enhancer (Westin et al, 1984;Ares et al, 1985;Mangin et al, 1986; for review see Dahlberg and Lund, 1987).…”

Cooperative interactions between transcription factors Sp1 and OTF-1.

“…U2 snRNA'genes are transcribed by RNA polymerase II (4,5), and the transcription is dependent on a distal enhancer that functions in conjunction with a proximal sequence element (PSE), located 50-60 base pairs (bp) upstream of the cap site (6)(7)(8)(9)(10). It has been shown (11,12) that the U2 enhancer includes one so called octamer motif ATGCAAAT (13), which is the binding site for the transcription factor OTF-1 in nonlymphoid cells (14), and three binding sites for the ubiquitous transcription factor Spl (15,16). A previous characterization of the U2 enhancer revealed a nonadditive functional cooperation between the OTF-1-and the adjacent Spl-binding sites (17).…”