title: Mapping the unique activation function 3 in the progesterone B-receptor upstream segment. Two LXXLL motifs and a tryptophan residue are required for activity.
abstract: Progesterone receptors (PR) contain three activation functions (AFs) that together define the extent to which they regulate transcription. AF1 and AF2 are common to the two isoforms of PR, PR-A and PR-B, whereas AF3 lies within the N-terminal 164 amino acids unique to PR-B, termed the "B-upstream segment" (BUS). To define the BUS regions that contribute to AF3 function, we generated a series of deletion and amino acid substitution mutants and tested them in three backgrounds as follows: BUS alone fused to the PR DNA binding domain (BUS-DBD), the entire PR-B N terminus linked to its DBD (NT-B), and full-length PR-B. Analyses of these mutants identified two regions in BUS whose loss reduces AF3 activity by more than 90%. These are associated with amino acids 54-90 (R1) and 120-154 (R2). R1 contains a consensus (55)LXXLL(59) motif (L1) identical to ones found in nuclear receptor co-activators. R2 is adjacent to a second nuclear receptor box (L2) at (115)LXXLL(119) and contains a conserved tryptophan (Trp-140). Their mutation completely disrupts AF3 activity in a promoter and cell type-independent manner. Critical mutations elicited similar effects on all three B-receptor backgrounds. This underscores the probability that these mutations alter a process linking BUS structure to the function of full-length PR-B in a fundamental way.
authors.affiliation: Department of Medicine, Division of Endocrinology, University of Colorado Health Sciences Center, 4200 East 9th Ave., Denver, CO 80262, USA.
authors.forename: L
authors.initials: L
authors.lastname: Tung
authors.forename: T
authors.initials: T
authors.lastname: Shen
authors.forename: M G
authors.initials: MG
authors.lastname: Abel
authors.forename: R L
authors.initials: RL
authors.lastname: Powell
authors.forename: G S
authors.initials: GS
authors.lastname: Takimoto
authors.forename: C A
authors.initials: CA
authors.lastname: Sartorius
authors.forename: K B
authors.initials: KB
authors.lastname: Horwitz
mesh_terms: D020816:Amino Acid Motifs; D001665:Binding Sites; D006801:Humans; D008565:Membrane Proteins; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010446:Peptide Fragments; D011980:Receptors, Progesterone / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins; D017384:Sequence Deletion; D014157:Transcription Factors / Q000378:metabolism*; D014158:Transcription, Genetic
chemical_list: C438144:ASI1 protein, S cerevisiae; D008565:Membrane Proteins; D010446:Peptide Fragments; D011980:Receptors, Progesterone; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; C097314:progesterone receptor B