scholarly journals An AU-rich element in the 3' untranslated region of the spinach chloroplast petD gene participates in sequence-specific RNA-protein complex formation.

1995 ◽  
Vol 15 (4) ◽  
pp. 2010-2018 ◽  
Author(s):  
Q Chen ◽  
C C Adams ◽  
L Usack ◽  
J Yang ◽  
R A Monde ◽  
...  
Keyword(s):  
Complex Formation ◽  
Protein Complex ◽  
Untranslated Region ◽  
Rna Binding ◽  
Rbcl Gene ◽  
Spinach Chloroplast ◽  
Protein Complex Formation ◽  
Sequence Elements ◽  
Chloroplast Mrna

In chloroplasts, the 3' untranslated regions of most mRNAs contain a stem-loop-forming inverted repeat (IR) sequence that is required for mRNA stability and correct 3'-end formation. The IR regions of several mRNAs are also known to bind chloroplast proteins, as judged from in vitro gel mobility shift and UV cross-linking assays, and these RNA-protein interactions may be involved in the regulation of chloroplast mRNA processing and/or stability. Here we describe in detail the RNA and protein components that are involved in 3' IR-containing RNA (3' IR-RNA)-protein complex formation for the spinach chloroplast petD gene, which encodes subunit IV of the cytochrome b6/f complex. We show that the complex contains 55-, 41-, and 29-kDa RNA-binding proteins (ribonucleoproteins [RNPs]). These proteins together protect a 90-nucleotide segment of RNA from RNase T1 digestion; this RNA contains the IR and downstream flanking sequences. Competition experiments using 3' IR-RNAs from the psbA or rbcL gene demonstrate that the RNPs have a strong specificity for the petD sequence. Site-directed mutagenesis was carried out to define the RNA sequence elements required for complex formation. These studies identified an 8-nucleotide AU-rich sequence downstream of the IR; mutations within this sequence had moderate to severe effects on RNA-protein complex formation. Although other similar sequences are present in the petD 3' untranslated region, only a single copy, which we have termed box II, appears to be essential for in vitro protein binding. In addition, the IR itself is necessary for optimal complex formation. These two sequence elements together with an RNP complex may direct correct 3'-end processing and/or influence the stability of petD mRNA in chloroplasts.

Novel Procedure To Investigate the Effect of Phosphorylation on Protein Complex Formation in Vitro and in Cells†

Biochemistry ◽  
2008 ◽  
Vol 47 (7) ◽  
pp. 2153-2161 ◽  
Author(s):  
Makoto Rembutsu ◽  
Marc P. M. Soutar ◽  
Lidy Van Aalten ◽  
Robert Gourlay ◽  
C. James Hastie ◽  
...  
Keyword(s):  
Complex Formation ◽  
Protein Complex ◽  
Protein Complex Formation ◽  
In Cells

BIOLOGIC AND BIOCHEMICAL EFFECTS OF ANTI-ANDROGENS ON RAT VENTRAL PROSTATE

1974 ◽  
Vol 75 (2) ◽  
pp. 385-397 ◽  
Author(s):  
Jack Geller ◽  
Kevin McCoy
Keyword(s):  
Complex Formation ◽  
Protein Complex ◽  
Ventral Prostate ◽  
Common Denominator ◽  
Biochemical Effects ◽  
Protein Complex Formation ◽  
Biologic Effects ◽  
Rat Prostate

ABSTRACT To determine whether the similarity of biologic effects of two antiandrogens, cyproterone acetate (Cyp A) and edogesterone (PH-218), could be related to one or more common biochemical effects, we have compared the effects of both drugs on 3H testosterone (3HT) entry into cells, binding to specific cytosol and nuclear androphiles, and conversion to dihydrotestosterone (DHT). In chronic in vivo studies, both Cyp A and PH-218 reduced rat prostate weights by approximately 50% and specific cytosol steroid-protein complex formation by approximately 60 %. At the same time, Cyp A decreased the formation of nuclear steroid-protein complex to 10% of control values, compared with 40% for PH-218. In addition, Cyp A, but not PH-218, significantly decreased total 3HT uptake by the prostate. Similar effects of Cyp A on 3HT uptake, binding, and metabolism were noted in acute in vivo and in vitro experiments. PH-218 effects on these same parameters were reduced in acute, compared to chronic, studies. Neither drug significantly affected the conversion of T to DHT. Despite quantitative differences between Cyp A and PH-218, these studies support the concept that the biochemical common denominator for the biologic effects of anti-androgens is inhibition of specific steroid-protein complex formation in both cytosol and nucleus.

scholarly journals cis- and trans-Acting Determinants for Translation of psbD mRNA inChlamydomonas reinhardtii

2000 ◽  
Vol 20 (21) ◽  
pp. 8134-8142 ◽  
Author(s):  
Friedrich Ossenbühl ◽  
Jörg Nickelsen
Keyword(s):  
Protein Complex ◽  
Rna Binding ◽  
Binding Protein ◽  
Rna Binding Protein ◽  
Clear Correlation ◽  
Protein Complex Formation ◽  
Chloroplast Translation ◽  
Rna Stabilization ◽  
Cis And Trans

ABSTRACT Chloroplast translation is mediated by nucleus-encoded factors that interact with distinct cis-acting RNA elements. A U-rich sequence within the 5′ untranslated region of the psbD mRNA has previously been shown to be required for its translation inChlamydomonas reinhardtii. By using UV cross-linking assays, we have identified a 40-kDa RNA binding protein, which binds to the wild-type psbD leader, but is unable to recognize a nonfunctional leader mutant lacking the U-rich motif. RNA binding is restored in a chloroplast cis-acting suppressor. The functions of several site-directed psbD leader mutants were analyzed with transgenic C. reinhardtii chloroplasts and the in vitro RNA binding assay. A clear correlation between photosynthetic activity and the capability to bind RNA by the 40-kDa protein was observed. Furthermore, the data obtained suggest that the poly(U) region serves as a molecular spacer between two previously characterized cis-acting elements, which are involved in RNA stabilization and translation. RNA-protein complex formation depends on the nuclear Nac2 gene product that is part of a protein complex required for the stabilization of the psbDmRNA. The sedimentation properties of the 40-kDa RNA binding protein suggest that it interacts directly with this Nac2 complex and, as a result, links processes of chloroplast RNA metabolism and translation.

scholarly journals Mutational Analysis of the Rhizobium etli recA Operator

1998 ◽  
Vol 180 (23) ◽  
pp. 6325-6331 ◽  
Author(s):  
Angels Tapias ◽  
Jordi Barbé
Keyword(s):  
Complex Formation ◽  
Protein Complex ◽  
Mutational Analysis ◽  
Rhizobium Meliloti ◽  
Regulatory Sequence ◽  
Rhizobium Etli ◽  
Wild Type ◽  
Protein Complex Formation

ABSTRACT Based upon our earlier studies (A. Tapias, A. R. Fernández de Henestrosa, and J. Barbé, J. Bacteriol. 179:1573–1579, 1997) we hypothesized that the regulatory sequence of the Rhizobium etli recA gene was TTGN11CAA. However, further detailed analysis of the R. etli recAoperator described in the present work suggests that it may in fact be GAACN7GTAC. This new conclusion is based upon PCR mutagenesis analysis carried out in the R. etli recAoperator, which indicates that the GAAC and GTAC submotifs found in the sequence GAACN7GTAC are required for the maximal stimulation of in vivo transcription and in vitro DNA-protein complex formation. This DNA-protein complex is also detected when the GAACN7GTAC wild-type sequence is modified to obtain GAACN7GAAC, GTACN7GTAC, or GAACN7GTTC. The wild-type promoters of the Rhizobium meliloti and Agrobacterium tumefaciens recA genes, which also contain the GAACN7GTAC sequence, compete with the R. etli recA promoter for the DNA-protein complex formation but not with mutant derivatives in any of these motifs, indicating that the R. etli, R. meliloti, andA. tumefaciens recA genes present the same regulatory sequence.

scholarly journals Analysis of Protein Complex Formation on Membrane Surfaces by Single Molecule Diffusion

2012 ◽  
Vol 102 (3) ◽  
pp. 183a
Author(s):  
Brian P. Ziemba ◽  
Jefferson D. Knight ◽  
Joseph J. Falke
Keyword(s):  
Complex Formation ◽  
Single Molecule ◽  
Protein Complex ◽  
Membrane Surfaces ◽  
Protein Complex Formation

Specific binding of chloroplast proteins in vitro to the 3' untranslated region of spinach chloroplast petD mRNA

1991 ◽  
Vol 11 (9) ◽  
pp. 4380-4388
Author(s):  
H C Chen ◽  
D B Stern
Keyword(s):  
Untranslated Region ◽  
Specific Binding ◽  
Rna Stability ◽  
Repeat Sequence ◽  
Hairpin Structure ◽  
Sequence Motif ◽  
Spinach Chloroplast ◽  
Higher Plant ◽  
Chloroplast Proteins

A detailed analysis of RNA-protein complex formation in the 3' untranslated region of spinach chloroplast petD mRNA has been carried out. Five chloroplast proteins that interact with petD RNA in this region, which contains an inverted repeat sequence capable of forming a hairpin structure, have been identified. A 33-kDa protein recognizes specifically the double-stranded stem of the hairpin structure; mutations that disrupt base pairing at the base of the stem reduce or eliminate protein binding. A 57-kDa protein recognizes specifically an AU-rich sequence motif that is highly conserved in petD genes of different higher plant species. The 57-kDa protein and possibly the 33-kDa protein form stable complexes with petD RNA in vitro and may interact with each other. In addition, their interaction with petD RNA is highly sensitive to heparin. The three other proteins, of 100, 32, and 28 kDa, display little sequence or structural binding specificity apart from their preference for uridine-rich sequences. They also interact with the 3' untranslated regions of other chloroplast RNAs such as those of psbA and rbcL. The functions of these proteins in the regulation of petD gene expression, including possible roles in transcription termination and RNA stability, are discussed.

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