scholarly journals Construction and Use of Derivatives of Transposon Tn4001 That Function in Mycoplasma pulmonis andMycoplasma arthritidis

2000 ◽  
Vol 182 (15) ◽  
pp. 4343-4347 ◽  
Author(s):  
Kevin Dybvig ◽  
C. Todd French ◽  
LeRoy L. Voelker
Keyword(s):  
Tetracycline Resistance ◽  
Chloramphenicol Acetyltransferase ◽  
Resistance Determinant ◽  
Mycoplasma Pulmonis ◽  
Functional Failure ◽  
General Utility ◽  
Lacz Activity ◽  
Gentamicin Resistance ◽  
Chloramphenicol Acetyltransferase Gene ◽  
Derivatives Of

ABSTRACT Previous attempts to introduce transposon Tn4001 intoMycoplasma pulmonis and Mycoplasma arthritidishave not been successful, possibly due to functional failure of the transposon's gentamicin resistance determinant. Tn4001Cand Tn4001T were constructed, respectively, by insertion of a chloramphenicol acetyltransferase gene and the tetMtetracycline resistance determinant into Tn4001. Both Tn4001C and Tn4001T transposed in M. pulmonis, and Tn4001T transposed in M. arthritidis. The incorporation of a Tn4001Tderivative that contained lacZ into eitherMycoplasma species resulted in transformants with readily detectable LacZ activity. Tn4001T may be of general utility for use as a mycoplasma cloning vehicle because tetMfunctions in all species of Mycoplasma examined thus far.

scholarly journals A direct role for C/EBP and the AP-I-binding site in gene expression linked to adipocyte differentiation.

1989 ◽  
Vol 9 (12) ◽  
pp. 5331-5339 ◽  
Author(s):  
R Herrera ◽  
H S Ro ◽  
G S Robinson ◽  
K G Xanthopoulos ◽  
B M Spiegelman
Keyword(s):  
Gene Expression ◽  
Transcriptional Activation ◽  
Adipocyte Differentiation ◽  
Binding Protein ◽  
Point Mutations ◽  
Chloramphenicol Acetyltransferase ◽  
Lipid Binding ◽  
Direct Role ◽  
New Genes ◽  
Chloramphenicol Acetyltransferase Gene

Adipocyte differentiation is accompanied by the transcriptional activation of many new genes, including the gene encoding adipocyte P2 (aP2), an intracellular lipid-binding protein. Using specific deletions and point mutations, we have shown that at least two distinct sequence elements in the aP2 promoter contribute to the expression of the chloramphenicol acetyltransferase gene in chimeric constructions transfected into adipose cells. An AP-I site at -120, shown earlier to bind Jun- and Fos-like proteins, serves as a positive regulator of chloramphenicol acetyltransferase gene expression in adipocytes but is specifically silenced by adjacent upstream sequences in preadipocytes. Sequences upstream of the AP-I site at -140 (termed AE-1) can function as an enhancer in both cell types when linked to a viral promoter but can stimulate expression only in fat cells in the intact aP2 promoter. The AE-1 sequence binds an adipocyte protein identical or very closely related to an enhancer-binding protein (C/EBP) that has been previously implicated in the regulation of several liver-specific genes. A functional role for C/EBP in the regulation of the aP2 gene is indicated by the facts that C/EBP mRNA is induced during adipocyte differentiation and the aP2 promoter is transactivated by cotransfection of a C/EBP expression vector into preadipose cells. These results indicate that sequences that bind C/EBP and the Fos-Jun complex play major roles in the expression of the aP2 gene during adipocyte differentiation and demonstrate that C/EBP can directly regulate cellular gene expression.

Generation of infectious Moloney murine leukemia viruses with deletions in the U3 portion of the long terminal repeat

1986 ◽  
Vol 6 (12) ◽  
pp. 4634-4640
Author(s):  
R Hanecak ◽  
S Mittal ◽  
B R Davis ◽  
H Fan
Keyword(s):  
Long Terminal Repeat ◽  
Transient Expression ◽  
Leukemia Virus ◽  
Chloramphenicol Acetyltransferase ◽  
Terminal Repeat ◽  
Tata Box ◽  
Moloney Murine Leukemia Virus ◽  
Chloramphenicol Acetyltransferase Gene ◽  
Leukemia Viruses ◽  
Murine Leukemia

Deletional analysis within the long terminal repeat (LTR) of Moloney murine leukemia virus (M-MuLV) was performed. By molecular cloning, deletions were made in the vicinity of the XbaI site at -150 base pairs (bp) in the U3 region, between the tandemly repeated enhancers and the TATA box. The effects of the deletions on LTR function were measured in two ways. First, deleted LTRs were fused to the bacterial chloramphenicol acetyltransferase gene and used in transient expression assays. Second, infectious M-MuLVs were generated by transfection of M-MuLV proviruses containing the deleted LTRs, and the relative infectivity of the mutant viruses was assessed by XC-syncytial assay. Most of the deleted LTRs examined showed relatively high promoter activity in the transient chloramphenicol acetyltransferase assays, with values ranging from 20 to 50% of the wild-type M-MuLV LTR. Thus, the sequences between the enhancers and the TATA box were not absolutely required for transient expression. However, infectivity of viruses carrying the same deleted LTRs showed more pronounced effects. Deletion of sequences from -195 to -174 bp reduced infectivity 20- to 100-fold. Deletion of sequences within the region from -174 to -122 bp did not affect infectivity, indicating that this region is dispensable. On the other hand, deletion of sequences from -150 to -40 bp reduced infectivity from 5 to 6 logs, although the magnitude of the reduction partly may have reflected threshold envelope protein requirements for positive XC assays. The reduced infectivity did not appear to result from a failure of proviral DNA synthesis or integration by the mutant. Thus, the infectivity measurements identified three functional domains in the region between the enhancers and the TATA box.

scholarly journals Novel Tetracycline Resistance Determinant from the Oral Metagenome

2003 ◽  
Vol 47 (4) ◽  
pp. 1430-1432 ◽  
Author(s):  
M. L. Diaz-Torres ◽  
R. McNab ◽  
D. A. Spratt ◽  
A. Villedieu ◽  
N. Hunt ◽  
...  
Keyword(s):  
Resistance Gene ◽  
Tetracycline Resistance ◽  
Resistance Determinant ◽  
Tetracycline Resistance Gene ◽  
Major Drawback

ABSTRACT A major drawback of most studies on how bacteria become resistant to antibiotics is that they concentrate mainly on bacteria that can be cultivated in the laboratory. In the present study, we cloned part of the oral metagenome and isolated a novel tetracycline resistance gene, tet(37), which inactivates tetracycline.

scholarly journals Novel Tetracycline Resistance Determinant Isolated from an Environmental Strain of Serratia marcescens

2007 ◽  
Vol 73 (7) ◽  
pp. 2199-2206 ◽  
Author(s):  
Stuart A. Thompson ◽  
Elizabeth V. Maani ◽  
Angela H. Lindell ◽  
Catherine J. King ◽  
J. Vaun McArthur
Keyword(s):  
Serratia Marcescens ◽  
Efflux Pump ◽  
Tetracycline Resistance ◽  
Amino Acid Identity ◽  
Resistance Determinant ◽  
Mercury Resistance ◽  
Resistance Locus ◽  
Environmental Strain ◽  
Apparent Lack ◽  
E Coli

ABSTRACT Resistances to tetracycline and mercury were identified in an environmental strain of Serratia marcescens isolated from a stream highly contaminated with heavy metals. As a step toward addressing the mechanisms of coselection of heavy metal and antibiotic resistances, the tetracycline resistance determinant was cloned in Escherichia coli. Within the cloned 13-kb segment, the tetracycline resistance locus was localized by deletion analysis and transposon mutagenesis. DNA sequence analysis of an 8.0-kb region revealed a novel gene [tetA(41)] that was predicted to encode a tetracycline efflux pump. Phylogenetic analysis showed that the TetA(41) protein was most closely related to the Tet(39) efflux protein of Acinetobacter spp. yet had less than 80% amino acid identity with known tetracycline efflux pumps. Adjacent to the tetA(41) gene was a divergently transcribed gene [tetR(41)] predicted to encode a tetracycline-responsive repressor protein. The tetA(41)-tetR(41) intergenic region contained putative operators for TetR(41) binding. The tetA(41) and tetR(41) promoters were analyzed using lacZ fusions, which showed that the expression of both the tetA(41) and tetR(41) genes exhibited TetR(41)-dependent regulation by subinhibitory concentrations of tetracycline. The apparent lack of plasmids in this S. marcescens strain, as well as the presence of metabolic genes adjacent to the tetracycline resistance locus, suggested that the genes were located on the S. marcescens chromosome and may have been acquired by transduction. The cloned Tet 41 determinant did not confer mercury resistance to E. coli, confirming that Tet 41 is a tetracycline-specific efflux pump rather than a multidrug transporter.

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