Oct2 transcription factors of the catfish (Ictalurus punctatus) are expressed as alternatively spliced alpha and beta isoforms. Functional analysis revealed an N-terminal glutamine (Q)-rich transactivation domain common to both isoforms of catfish Oct2. A C-terminal proline, serine, threonine (PST)-rich activation domain was identified exclusively in the beta isoform. Activation domains of fish and mammalian Oct2 showed cell type- and species-specific activity correlated with their biochemical composition (Q-rich vs PST-rich). In contrast the activation domains of the aryl hydrocarbon receptor and aryl hydrocarbon receptor nuclear translocator of fish and mammals showed no correlation of activity with biochemical composition or species of origin. Although isolated catfish Oct2 activation domains were unable to drive transcription from a site 1.9kb distal to the promoter, Oct2beta activated transcription from both an IgH enhancer and an array of octamer motifs at this distal position. The properties of catfish Oct2 activation domains differ depending on whether they are studied in isolation or as components of the intact transcription factor.
"Fu and Li's D statistic was significantly positive (at a p-value of 0.01) in the New Bedford Harbor population at the sliding windows of nucleotide regions 1249 to 1340 bp. This region contains a predicted Oct2 transcription factor binding site, an enhancer that has been found to regulate immunoglobulin heavy-chain (IgH) genes in other teleosts (Cioffi et al., 2002). "
[Show abstract][Hide abstract] ABSTRACT: The functional importance of variable, transcriptional regulatory sequences within and among natural populations is largely unexplored. We analysed the cytochrome P4501A (CYP1A) promoter in three populations of the minnow, Fundulus heteroclitus, because two SNPs in the promoter and first intron of CYP1A are under selection in populations adapted to pollutants. To define the importance of these SNPs, 1630 bp of the CYP1A promoter and first intron and exon were sequenced in eight individuals from three populations: a population from a polluted environment resistant to some aromatic pollutants and two flanking reference populations. CYP1A is induced by many aromatic pollutants, but in populations adapted to pollutants, CYP1A has been shown to be refractory to induction. We were interested in understanding whether variation in the CYP1A promoter explains mechanism(s) of adaptation to these aromatic pollutants. The CYP1A promoter was extremely variable (an average of 9.3% of the promoter nucleotides varied among all populations) and exhibited no fixed differences between populations. As CYP1A is poorly inducible in adapted fish, we hypothesized that CYP1A promoter regions might vary functionally between populations. Unexpectedly, in vitro analysis showed significantly greater transcription from CYP1A promoters found in the population from the polluted environment relative to promoters found in both reference populations. Thus, despite extensive variation among populations and lack of fixed differences between populations, individuals from a polluted environment have significantly enhanced promoter activity. These data demonstrate that intraspecific variation, which provides the raw material for natural selection to act on, can occur while maintaining promoter function.
"ons that encode transcrip - tion factor domains that bind either DNA motifs or other protein domains . Finally , it is evident from the published literature that knowledge of the pres - ence and expression of a transcription factor gene in teleost fish is insufficient evidence on which to base assumptions about its function ( Ross et al . , 1998 ; Cioffi et al . , 2002 ; Hikima et al . , 2004 , 2005b , 2006a , 2006b ; Lennard et al . , 2007 ) ."
[Show abstract][Hide abstract] ABSTRACT: The immunoglobulin heavy chain locus of teleost fish is driven by an enhancer (Eμ3′) that contains, in all species examined, octamer and E-box motifs (typically μE5). Thus, the transcription factors known (through experiments in fish) or predicted (from knowledge of their mammalian homologues) to be involved in the function of the enhancer are the octamer-binding (Oct) transcription factors Oct1 and Oct2, their coactivator BOB.1 and the μE5-binding E-proteins (E2A, HEB and E2-2). Comparative genomic analysis of the genes encoding these transcription factors in teleost fish permits some conclusions to be drawn concerning the fate of these genes following the genome-wide duplication event that occurred in this lineage. Although problems in the assembly of some regions of the zebrafish genome may complicate the analysis, the pattern that emerges from these studies is that Oct1, Oct2, BOB.1 and E2-2 are all single-copy genes in teleosts, whereas in the case of E2A and HEB, two copies of each gene are present. The evolutionary conservation of sequences between related genes is variable, but with the highest similarities always being observed in the domains with ligand-binding function, whether the ligand is a DNA motif or, as in the case of BOB.1, another protein domain. A major challenge that must be faced is the assignment of precise functions to these diverse transcription factors in teleosts; assumptions drawn by inference from their mammalian homologues will likely be misleading.
Journal of Fish Biology 09/2007; 71(sb):153 - 173. DOI:10.1111/j.1095-8649.2007.01632.x · 1.66 Impact Factor
"This construct contains 5 Gal4 binding sites, located upstream of a minimal TATA box promoter and the CAT reporter gene. The expression constructs for the fusion proteins containing the Gal4 DNA binding domain (DBD) alone or with VP16; nucleolin; catfish Oct2β N-terminus; and catfish Oct2β C-terminus have previously been described . The Gal4DBD-Oct1 domain fusion expression constructs were produced by PCR and were directionally cloned into the Gal4DBD expression construct with XbaI and BamHI restriction enzyme sites. "
[Show abstract][Hide abstract] ABSTRACT: The enhancer (Emu3') of the immunoglobulin heavy chain locus (IGH) of the channel catfish (Ictalurus punctatus) has been well characterized. The functional core region consists of two variant Oct transcription factor binding octamer motifs and one E-protein binding muE5 site. An orthologue to the Oct2 transcription factor has previously been cloned in catfish and is a functionally active transcription factor. This study was undertaken to clone and characterize the Oct1 transcription factor, which has also been shown to be important in driving immunoglobulin gene transcription in mammals.
An orthologue of Oct1, a POU family transcription factor, was cloned from a catfish macrophage cDNA library. The inferred amino acid sequence of the catfish Oct1, when aligned with other vertebrate Oct1 sequences, revealed clear conservation of structure, with the POU specific subdomain of catfish Oct1 showing 96% identity to that of mouse Oct1. Expression of Oct1 was observed in clonal T and B cell lines and in all tissues examined. Catfish Oct1, when transfected into both mammalian (mouse) and catfish B cell lines, unexpectedly failed to drive transcription from three different octamer-containing reporter constructs. These contained a trimer of octamer motifs, a fish VH promoter, and the core region of the catfish Emu3' IGH enhancer, respectively. This failure of catfish Oct1 to drive transcription was not rescued by human BOB.1, a co-activator of Oct transcription factors that stimulates transcription driven by catfish Oct2. When co-transfected with catfish Oct2, Oct1 reduced Oct2 driven transcriptional activation. Electrophoretic mobility shift assays showed that catfish Oct1 (native or expressed in vitro) bound both consensus and variant octamer motifs. Putative N- and C-terminal activation domains of Oct1, when fused to a Gal4 DNA binding domain and co-transfected with Gal4-dependent reporter constructs were transcriptionally inactive, which may be due in part to a lack of residues associated with activation domain function.
An orthologue to mammalian Oct1 has been found in the catfish. It is similar to mammalian Oct1 in structure and expression. However, these results indicate that the physiological functions of catfish Oct1 differ from those of mammalian Oct1 and include negative regulation of transcription.
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