Expression of a yeast metallothionein gene family is activated by a single metalloregulatory transcription factor

1992 ◽  
Vol 12 (9) ◽  
pp. 3766-3775
Author(s):  
P Zhou ◽  
M S Szczypka ◽  
T Sosinowski ◽  
D J Thiele
Keyword(s):  
Gene Family ◽  
Protein Complexes ◽  
Copper Resistance ◽  
Yeast Saccharomyces Cerevisiae ◽  
Pathogenic Yeast ◽  
Selection Scheme ◽  
Metallothionein Gene ◽  
Metal Levels ◽  
Opportunistic Pathogenic

The opportunistic pathogenic yeast Candida glabrata elicits at least two major responses in the presence of high environmental metal levels: transcriptional induction of the metallothionein gene family by copper and the appearance of small (gamma-Glu-Cys)nGly peptides in the presence of cadmium. On the basis of a trans-activation selection scheme in the baker's yeast Saccharomyces cerevisiae, we previously isolated a C. glabrata gene which encodes a copper-activated DNA-binding protein designated AMT1. AMT1 forms multiple specific DNA-protein complexes with both C. glabrata MT-I and MT-IIa promoter DNA fragments. In this report, we localize and define the AMT1-binding sites in the MT-I and MT-IIa promoters and characterize the mode of AMT1 binding. Furthermore, we demonstrate that the AMT1 protein trans activates both the MT-I and MT-IIa genes in vivo in response to copper and that this activation is essential for high-level copper resistance in C. glabrata. Although AMT1-mediated trans activation of the C. glabrata metallothionein genes is essential for copper resistance, AMT1 is completely dispensable for cadmium tolerance. The distinct function that metallothionein genes have in copper but not cadmium detoxification in C. glabrata is in contrast to the role that metallothionein genes play in tolerance to multiple metals in higher organisms.

scholarly journals Expression of a yeast metallothionein gene family is activated by a single metalloregulatory transcription factor.

1992 ◽  
Vol 12 (9) ◽  
pp. 3766-3775 ◽  
Author(s):  
P Zhou ◽  
M S Szczypka ◽  
T Sosinowski ◽  
D J Thiele
Keyword(s):  
Gene Family ◽  
Protein Complexes ◽  
Copper Resistance ◽  
Yeast Saccharomyces Cerevisiae ◽  
Pathogenic Yeast ◽  
Selection Scheme ◽  
Metallothionein Gene ◽  
Metal Levels ◽  
Opportunistic Pathogenic

The opportunistic pathogenic yeast Candida glabrata elicits at least two major responses in the presence of high environmental metal levels: transcriptional induction of the metallothionein gene family by copper and the appearance of small (gamma-Glu-Cys)nGly peptides in the presence of cadmium. On the basis of a trans-activation selection scheme in the baker's yeast Saccharomyces cerevisiae, we previously isolated a C. glabrata gene which encodes a copper-activated DNA-binding protein designated AMT1. AMT1 forms multiple specific DNA-protein complexes with both C. glabrata MT-I and MT-IIa promoter DNA fragments. In this report, we localize and define the AMT1-binding sites in the MT-I and MT-IIa promoters and characterize the mode of AMT1 binding. Furthermore, we demonstrate that the AMT1 protein trans activates both the MT-I and MT-IIa genes in vivo in response to copper and that this activation is essential for high-level copper resistance in C. glabrata. Although AMT1-mediated trans activation of the C. glabrata metallothionein genes is essential for copper resistance, AMT1 is completely dispensable for cadmium tolerance. The distinct function that metallothionein genes have in copper but not cadmium detoxification in C. glabrata is in contrast to the role that metallothionein genes play in tolerance to multiple metals in higher organisms.

scholarly journals A Family Knockout of All Four Drosophila Metallothioneins Reveals a Central Role in Copper Homeostasis and Detoxification

2006 ◽  
Vol 26 (6) ◽  
pp. 2286-2296 ◽  
Author(s):  
Dieter Egli ◽  
Hasmik Yepiskoposyan ◽  
Anand Selvaraj ◽  
Kuppusamy Balamurugan ◽  
Rama Rajaram ◽  
...  
Keyword(s):  
Gene Family ◽  
Tissue Specificity ◽  
Biochemical Characterization ◽  
Adult Males ◽  
Metallothionein Gene ◽  
Copper Cells ◽  
Complex Forms ◽  
Main Regulator ◽  
Distribution In The Organism

ABSTRACT Metallothioneins are ubiquitous, small, cysteine-rich proteins with the ability to bind heavy metals. In spite of their biochemical characterization, their in vivo function remains elusive. Here, we report the generation of a metallothionein gene family knockout in Drosophila melanogaster by targeted disruption of all four genes (MtnA to -D). These flies are viable if raised in standard laboratory food. During development, however, they are highly sensitive to copper, cadmium, and (to a lesser extent) zinc load. Metallothionein expression is particularly important for male viability; while copper load during development affects males and females equally, adult males lacking metallothioneins display a severely reduced life span, possibly due to copper-mediated oxidative stress. Using various reporter gene constructs, we find that different metallothioneins are expressed with virtually the same tissue specificity in larvae, notably in the intestinal tract at sites of metal accumulation, including the midgut's “copper cells.” The same expression pattern is observed with a synthetic minipromoter consisting only of four tandem metal response elements. From these and other experiments, we conclude that tissue specificity of metallothionein expression is a consequence, rather than a cause, of metal distribution in the organism. The bright orange luminescence of copper accumulated in copper cells of the midgut is severely reduced in the metallothionein gene family knockout, as well as in mutants of metal-responsive transcription factor 1 (MTF-1), the main regulator of metallothionein expression. This indicates that an in vivo metallothionein-copper complex forms the basis of this luminescence. Strikingly, metallothionein mutants show an increased, MTF-1-dependent induction of metallothionein promoters in response to copper, cadmium, silver, zinc, and mercury. We conclude that free metal, but not metallothionein-bound metal, triggers the activation of MTF-1 and that metallothioneins regulate their own expression by a negative feedback loop.

scholarly journals A novel 66-kilodalton protein complexes with Rrn6, Rrn7, and TATA-binding protein to promote polymerase I transcription initiation in Saccharomyces cerevisiae.

1996 ◽  
Vol 16 (11) ◽  
pp. 6436-6443 ◽  
Author(s):  
C W Lin ◽  
B Moorefield ◽  
J Payne ◽  
P Aprikian ◽  
K Mitomo ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Transcription Initiation ◽  
Protein Complexes ◽  
Binding Protein ◽  
Strong Association ◽  
Tata Binding Protein ◽  
Yeast Saccharomyces Cerevisiae ◽  
Pol I ◽  
Polymerase I

We report the cloning of RRN11, a gene coding for a 66-kDa protein essential for transcription initiation by RNA polymerase I (Pol I) in the yeast Saccharomyces cerevisiae. Rrn11 specifically complexes with two previously identified transcription factors, Rrn6 and Rrn7 (D. A. Keys, J. S. Steffan, J. A. Dodd, R. T. Yamamoto, Y. Nogi, and M. Nomura, Genes Dev. 8:2349-2362, 1994). The Rrn11-Rrn6-Rrn7 complex also binds the TATA-binding protein and is required for transcription by the core domain of the Pol I promoter. Therefore, we have designated the Rrn11-Rrn6-Rrn7-TATA-binding protein complex the yeast Pol I core factor. A two-hybrid assay was used to demonstrate involvement of short leucine heptad repeats on both Rrn11 and Rrn6 in the in vivo association of these two proteins. This assay also verified the previously described strong association between Rrn6 and Rrn7, independent of the Rrn6 leucine repeat.

Photoinhibition in vivo and in vitro Involves Weakly Coupled Chlorophyll–Protein Complexes‡¶

2002 ◽  
Vol 75 (6) ◽  
pp. 613 ◽  
Author(s):  
Stefano Santabarbara ◽  
Ilaria Cazzalini ◽  
Andrea Rivadossi ◽  
Flavio M. Garlaschi ◽  
Giuseppe Zucchelli ◽  
...  
Keyword(s):  
Protein Complexes ◽  
Weakly Coupled ◽  
Chlorophyll Protein Complexes ◽  
Chlorophyll Protein

scholarly journals RNA-Centric Approaches to Profile the RNA–Protein Interaction Landscape on Selected RNAs

2021 ◽  
Vol 7 (1) ◽  
pp. 11 ◽  
Author(s):  
André P. Gerber
Keyword(s):  
Mass Spectrometry ◽  
Protein Interactions ◽  
Regulatory Networks ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Protein Complexes ◽  
Cell Protein ◽  
Transcriptional Regulatory Networks ◽  
Technological Advances

RNA–protein interactions frame post-transcriptional regulatory networks and modulate transcription and epigenetics. While the technological advances in RNA sequencing have significantly expanded the repertoire of RNAs, recently developed biochemical approaches combined with sensitive mass-spectrometry have revealed hundreds of previously unrecognized and potentially novel RNA-binding proteins. Nevertheless, a major challenge remains to understand how the thousands of RNA molecules and their interacting proteins assemble and control the fate of each individual RNA in a cell. Here, I review recent methodological advances to approach this problem through systematic identification of proteins that interact with particular RNAs in living cells. Thereby, a specific focus is given to in vivo approaches that involve crosslinking of RNA–protein interactions through ultraviolet irradiation or treatment of cells with chemicals, followed by capture of the RNA under study with antisense-oligonucleotides and identification of bound proteins with mass-spectrometry. Several recent studies defining interactomes of long non-coding RNAs, viral RNAs, as well as mRNAs are highlighted, and short reference is given to recent in-cell protein labeling techniques. These recent experimental improvements could open the door for broader applications and to study the remodeling of RNA–protein complexes upon different environmental cues and in disease.

scholarly journals Cloning of mouse ptx3, a new member of the pentraxin gene family expressed at extrahepatic sites

Blood ◽  
1996 ◽  
Vol 87 (5) ◽  
pp. 1862-1872 ◽  
Author(s):  
M Introna ◽  
VV Alles ◽  
M Castellano ◽  
G Picardi ◽  
L De Gioia ◽  
...  
Keyword(s):  
Gene Family ◽  
Tertiary Structure ◽  
Helical Structure ◽  
Distinct Pattern ◽  
Mononuclear Phagocytes ◽  
Acute Phase Reactants ◽  
Inflammatory Reactions ◽  
Reactive Protein

Abstract Pentraxins, which include C reactive protein (CRP) and serum amyloid P component (SAP), are prototypic acute phase reactants that serve as indicators of inflammatory reactions. Here we report genomic and cDNA cloning of mouse ptx3 (mptx3), a member of the pentraxin gene family and characterize its extrahepatic expression in vitro and in vivo. mptx3 is organized into three exons on chromosome 3: the first (43 aa) and second exon (175 aa) code for the signal peptide and for a protein portion with no high similarity to known sequences the third (203 aa) for a domain related to classical pentraxins, which contains the “pentraxin family signature.” Analysis of the N terminal portion predicts a predominantly alpha helical structure, while the pentraxin domain of ptx3 is accommodated comfortably in the tertiary structure fold of SAP. Normal and transformed fibroblasts, undifferentiated and differentiated myoblasts, normal endothelial cells, and mononuclear phagocytes express mptx3 mRNA and release the protein in vitro on exposure to interleukin-1beta (IL-1beta) and tumor necrosis factor (TNF)alpha. mptx3 was induced by bacterial lipopolysaccharide in vivo in a variety of organs and, most strongly, in the vascular endothelium of skeletal muscle and heart. Thus, mptx3 shows a distinct pattern of in vivo expression indicative of a significant role in cardiovascular and inflammatory pathology.

Conformational Flexibility in Assembly of Photosynthetic Membrane Protein Complexes in vivo

2004 ◽  
Vol 10 (S02) ◽  
pp. 1496-1497
Author(s):  
P A Bullough
Keyword(s):  
Membrane Protein ◽  
Protein Complexes ◽  
Conformational Flexibility ◽  
Photosynthetic Membrane ◽  
Membrane Protein Complexes

Extended abstract of a paper presented at Microscopy and Microanalysis 2004 in Savannah, Georgia, USA, August 1–5, 2004.

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