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Category: Bacterial Pathogenesis
Regulation by Termination-Antitermination: a Genomic Approach, Page 1 of 2
< Previous page | Next page > /docserver/preview/fulltext/10.1128/9781555817992/9781555812058_Chap23-1.gif /docserver/preview/fulltext/10.1128/9781555817992/9781555812058_Chap23-2.gifAbstract:
A variety of transcription termination-antitermination regulatory processes, often called transcription attenuation mechanisms, have been discovered in phage and microbial species. This chapter discusses known and predicted examples of regulation by antitermination-termination at sites specifying an intrinsic terminator, deduced by examining the entire Bacillus subtilis genome. In many mechanisms of transcription attenuation, the termination decision is based on whether an RNA antiterminator (AT) structure has been previously formed. To identify the operons of B. subtilis that may be regulated by transcription termination-antitermination, a computer program that searches the genome for overlapping AT/terminator (T) elements was developed based on nucleotide sequence and free energy criteria. The program designed was used to search the entire genome sequence of B. subtilis for sequences that could specify transcript AAT, AT, and T structures. The number of gene sequences identified that were preceded by potential AT and T structures was dependent on the parameters employed in the program. Conservation of the specific transcription attenuation regulatory mechanisms used for many genes encoding aminoacyl-tRNA synthetases or amino acid biosynthetic enzymes of B. subtilis and other gram-positive organisms has been well documented by Grundy and Henkin.
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Characteristics of the presumed AT/T or AAT/AT/T regulatory regions preceding genes trpE (A), yqgN (B), pyrP (C), and ykrW (D). Sequences corresponding to the -35 and -10 elements of the putative sigma A promoter preceding each gene are indicated in outlined letters. Sequences corresponding to the T, AT, and AAT structures are underlined with a single solid line, a single wavy line, and double solid lines, respectively. Common bases in the AT and T structures are in boldface, and common bases in the AAT and AT structures are in boldface italics. A more complete description of these sequences and structures can be obtained from our website (http://cmgm.stanford.edu/∼merino/Bacillus_subtilis/index.html .
Characteristics of the presumed AT/T or AAT/AT/T regulatory regions preceding genes trpE (A), yqgN (B), pyrP (C), and ykrW (D). Sequences corresponding to the -35 and -10 elements of the putative sigma A promoter preceding each gene are indicated in outlined letters. Sequences corresponding to the T, AT, and AAT structures are underlined with a single solid line, a single wavy line, and double solid lines, respectively. Common bases in the AT and T structures are in boldface, and common bases in the AAT and AT structures are in boldface italics. A more complete description of these sequences and structures can be obtained from our website (http://cmgm.stanford.edu/∼merino/Bacillus_subtilis/index.html .
Characteristics of the presumed AT/T or AAT/AT/T regulatory regions preceding genes trpE (A), yqgN (B), pyrP (C), and ykrW (D). Sequences corresponding to the -35 and -10 elements of the putative sigma A promoter preceding each gene are indicated in outlined letters. Sequences corresponding to the T, AT, and AAT structures are underlined with a single solid line, a single wavy line, and double solid lines, respectively. Common bases in the AT and T structures are in boldface, and common bases in the AAT and AT structures are in boldface italics. A more complete description of these sequences and structures can be obtained from our website (http://cmgm.stanford.edu/∼merino/Bacillus_subtilis/index.html .
Characteristics of the presumed AT/T or AAT/AT/T regulatory regions preceding genes trpE (A), yqgN (B), pyrP (C), and ykrW (D). Sequences corresponding to the -35 and -10 elements of the putative sigma A promoter preceding each gene are indicated in outlined letters. Sequences corresponding to the T, AT, and AAT structures are underlined with a single solid line, a single wavy line, and double solid lines, respectively. Common bases in the AT and T structures are in boldface, and common bases in the AAT and AT structures are in boldface italics. A more complete description of these sequences and structures can be obtained from our website (http://cmgm.stanford.edu/∼merino/Bacillus_subtilis/index.html .
Characteristics of the presumed AT/T or AAT/AT/T regulatory regions preceding genes trpE (A), yqgN (B), pyrP (C), and ykrW (D). Sequences corresponding to the -35 and -10 elements of the putative sigma A promoter preceding each gene are indicated in outlined letters. Sequences corresponding to the T, AT, and AAT structures are underlined with a single solid line, a single wavy line, and double solid lines, respectively. Common bases in the AT and T structures are in boldface, and common bases in the AAT and AT structures are in boldface italics. A more complete description of these sequences and structures can be obtained from our website (http://cmgm.stanford.edu/∼merino/Bacillus_subtilis/index.html .
Characteristics of the presumed AT/T or AAT/AT/T regulatory regions preceding genes trpE (A), yqgN (B), pyrP (C), and ykrW (D). Sequences corresponding to the -35 and -10 elements of the putative sigma A promoter preceding each gene are indicated in outlined letters. Sequences corresponding to the T, AT, and AAT structures are underlined with a single solid line, a single wavy line, and double solid lines, respectively. Common bases in the AT and T structures are in boldface, and common bases in the AAT and AT structures are in boldface italics. A more complete description of these sequences and structures can be obtained from our website (http://cmgm.stanford.edu/∼merino/Bacillus_subtilis/index.html .
Characteristics of the presumed AT/T or AAT/AT/T regulatory regions preceding genes trpE (A), yqgN (B), pyrP (C), and ykrW (D). Sequences corresponding to the -35 and -10 elements of the putative sigma A promoter preceding each gene are indicated in outlined letters. Sequences corresponding to the T, AT, and AAT structures are underlined with a single solid line, a single wavy line, and double solid lines, respectively. Common bases in the AT and T structures are in boldface, and common bases in the AAT and AT structures are in boldface italics. A more complete description of these sequences and structures can be obtained from our website (http://cmgm.stanford.edu/∼merino/Bacillus_subtilis/index.html .
Characteristics of the presumed AT/T or AAT/AT/T regulatory regions preceding genes trpE (A), yqgN (B), pyrP (C), and ykrW (D). Sequences corresponding to the -35 and -10 elements of the putative sigma A promoter preceding each gene are indicated in outlined letters. Sequences corresponding to the T, AT, and AAT structures are underlined with a single solid line, a single wavy line, and double solid lines, respectively. Common bases in the AT and T structures are in boldface, and common bases in the AAT and AT structures are in boldface italics. A more complete description of these sequences and structures can be obtained from our website (http://cmgm.stanford.edu/∼merino/Bacillus_subtilis/index.html .
B. subtilis genes known or proposed to be regulated by antiterminator and terminator sequences and structures
a The homologous E. coli gene is also known to be regulated by attentuation.
b The homologous gene of E. coli was found by our program to have an attenuator.
c Gene regulated by the T-box mechanism ( 5 , 6 ).
d Gene regulated by the S-box mechanism ( 7 ).
e Gene regulated by TRAP (trp RNA-binding attenuation protein) ( 19 ).
f Direction of transcription of the upstream gene. S, same direction, D, opposite direction.
g Sequences that form antiterminator structures are in italics, and those that form terminator structures are in boldface.
h Coding sequences are in upper case, and intergenic sequences are in lower case.
i Based on GenBank sequence annotations, our program reads the upstream sequence of every gene. This sequence is considered as the analysis window and its default value is 300 nt. In this window, the program searches for a run of t's (rts) that would correspond to a run of u's in RNA. The size of the rts selected is 6 nt, at least five of which must be “ts”. tttatt or tttttc are examples of acceptable rts. The arbitrarily selected maximum acceptable space between the its and the first base of the downstream gene, is 110 nt. For every rts, the program looks for the most stable secondary structure that could be formed within the first upstream 60 nt. Initially, the secondary structure that could be formed from this 60 nt is predicted using the FoldRNA program ( 22 ). Trie only secondary structure that is considered part of a T is the stem-and'loop structure (SLS). If the sequence analyzed contains more than one SLS, only the one closest to the rts is considered in the analysis. In the event that the folding predictions yield structures different than an SLS (e.g., cloverleaf like), the first nt of the 60 nt long sequence is removed and the search for the SLS is repeated recursively until the program finds the sequence, which when folded, corresponds to the most stable SLS. This SLS is considered part of a T only if the base at the bottom of the stem is no more than 4 nt from the rts, the maximum number of loops in the structure is 3, and if the free energy of the structure is at least —0.2 kcal/mol per nucleotide.
g For every T found, the program then searches for the presence of an associated AT. The program begins by scanning a 60-nt sequence upstream of the middle of the main T loop. The program then finds the most stable SLS using the procedure described in the T analysis. An SLS is considered an AT on the basis of the following somewhat arbitrary preferences: If at least 3 bases of its stem overlap with the T sequence, the maximum number of loops of the structure is 4, and if it has a free energy that is at least one fourth that of its corresponding T. The previous analysis is also repeated on each AT, searching for its corresponding AAT structure. To consider the AAT and AT as mutually exclusive structures, they must share at least 3 bases. The free energy of the AAT must be at least one fourth that of its associated AT. Finally, the program looks for promoter sequences in the 300-bp upstream region of the AT/T elements using the algorithm proposed by Mulligan et al. ( 14 ). It also searches for some other regulatory sequence in the RNA leader region, such as the T-box sequence ( 5 , 6 ), or the TRAP binding site ( 18 , 19 ).
B. subtilis genes known or proposed to be regulated by antiterminator and terminator sequences and structures
a The homologous E. coli gene is also known to be regulated by attentuation.
b The homologous gene of E. coli was found by our program to have an attenuator.
c Gene regulated by the T-box mechanism ( 5 , 6 ).
d Gene regulated by the S-box mechanism ( 7 ).
e Gene regulated by TRAP (trp RNA-binding attenuation protein) ( 19 ).
f Direction of transcription of the upstream gene. S, same direction, D, opposite direction.
g Sequences that form antiterminator structures are in italics, and those that form terminator structures are in boldface.
h Coding sequences are in upper case, and intergenic sequences are in lower case.
i Based on GenBank sequence annotations, our program reads the upstream sequence of every gene. This sequence is considered as the analysis window and its default value is 300 nt. In this window, the program searches for a run of t's (rts) that would correspond to a run of u's in RNA. The size of the rts selected is 6 nt, at least five of which must be “ts”. tttatt or tttttc are examples of acceptable rts. The arbitrarily selected maximum acceptable space between the its and the first base of the downstream gene, is 110 nt. For every rts, the program looks for the most stable secondary structure that could be formed within the first upstream 60 nt. Initially, the secondary structure that could be formed from this 60 nt is predicted using the FoldRNA program ( 22 ). Trie only secondary structure that is considered part of a T is the stem-and'loop structure (SLS). If the sequence analyzed contains more than one SLS, only the one closest to the rts is considered in the analysis. In the event that the folding predictions yield structures different than an SLS (e.g., cloverleaf like), the first nt of the 60 nt long sequence is removed and the search for the SLS is repeated recursively until the program finds the sequence, which when folded, corresponds to the most stable SLS. This SLS is considered part of a T only if the base at the bottom of the stem is no more than 4 nt from the rts, the maximum number of loops in the structure is 3, and if the free energy of the structure is at least —0.2 kcal/mol per nucleotide.
g For every T found, the program then searches for the presence of an associated AT. The program begins by scanning a 60-nt sequence upstream of the middle of the main T loop. The program then finds the most stable SLS using the procedure described in the T analysis. An SLS is considered an AT on the basis of the following somewhat arbitrary preferences: If at least 3 bases of its stem overlap with the T sequence, the maximum number of loops of the structure is 4, and if it has a free energy that is at least one fourth that of its corresponding T. The previous analysis is also repeated on each AT, searching for its corresponding AAT structure. To consider the AAT and AT as mutually exclusive structures, they must share at least 3 bases. The free energy of the AAT must be at least one fourth that of its associated AT. Finally, the program looks for promoter sequences in the 300-bp upstream region of the AT/T elements using the algorithm proposed by Mulligan et al. ( 14 ). It also searches for some other regulatory sequence in the RNA leader region, such as the T-box sequence ( 5 , 6 ), or the TRAP binding site ( 18 , 19 ).
B. subtilis genes known or proposed to be regulated by anti-antiterminator, antiterminator, and terminator sequences and structures
a The homologous E. coli gene is also known to be regulated by attenuation.
b The homologous gene of E. coli was found by our program to have an attenuator.
c Gene regulated by the T-box mechanism ( 5 , 6 ).
d Gene regulated by the S-box mechanism ( 7 ).
e Gene regulated by TRAP (trp RNA–binding attenuation protein ( 19 ).
f Direction of transcription of the upstream gene. S, same direction, D, opposite direction.
g Sequences that form anti-antiterminator structures are underlined, those that form antiterminator structures are in italics, and those that form terminator structures are in boldface.
h Coding sequences are in upper case, and intergenic sequences are in lower case.
B. subtilis genes known or proposed to be regulated by anti-antiterminator, antiterminator, and terminator sequences and structures
a The homologous E. coli gene is also known to be regulated by attenuation.
b The homologous gene of E. coli was found by our program to have an attenuator.
c Gene regulated by the T-box mechanism ( 5 , 6 ).
d Gene regulated by the S-box mechanism ( 7 ).
e Gene regulated by TRAP (trp RNA–binding attenuation protein ( 19 ).
f Direction of transcription of the upstream gene. S, same direction, D, opposite direction.
g Sequences that form anti-antiterminator structures are underlined, those that form antiterminator structures are in italics, and those that form terminator structures are in boldface.
h Coding sequences are in upper case, and intergenic sequences are in lower case.
B. subtilis genes known or proposed to be regulated by anti-antiterminator, antiterminator, and terminator sequences and structures
a The homologous E. coli gene is also known to be regulated by attenuation.
b The homologous gene of E. coli was found by our program to have an attenuator.
c Gene regulated by the T-box mechanism ( 5 , 6 ).
d Gene regulated by the S-box mechanism ( 7 ).
e Gene regulated by TRAP (trp RNA–binding attenuation protein ( 19 ).
f Direction of transcription of the upstream gene. S, same direction, D, opposite direction.
g Sequences that form anti-antiterminator structures are underlined, those that form antiterminator structures are in italics, and those that form terminator structures are in boldface.
h Coding sequences are in upper case, and intergenic sequences are in lower case.
B. subtilis genes known or proposed to be regulated by anti-antiterminator, antiterminator, and terminator sequences and structures
a The homologous E. coli gene is also known to be regulated by attenuation.
b The homologous gene of E. coli was found by our program to have an attenuator.
c Gene regulated by the T-box mechanism ( 5 , 6 ).
d Gene regulated by the S-box mechanism ( 7 ).
e Gene regulated by TRAP (trp RNA–binding attenuation protein ( 19 ).
f Direction of transcription of the upstream gene. S, same direction, D, opposite direction.
g Sequences that form anti-antiterminator structures are underlined, those that form antiterminator structures are in italics, and those that form terminator structures are in boldface.
h Coding sequences are in upper case, and intergenic sequences are in lower case.
B. subtilis genes known or proposed to be regulated by anti-antiterminator, antiterminator, and terminator sequences and structures
a The homologous E. coli gene is also known to be regulated by attenuation.
b The homologous gene of E. coli was found by our program to have an attenuator.
c Gene regulated by the T-box mechanism ( 5 , 6 ).
d Gene regulated by the S-box mechanism ( 7 ).
e Gene regulated by TRAP (trp RNA–binding attenuation protein ( 19 ).
f Direction of transcription of the upstream gene. S, same direction, D, opposite direction.
g Sequences that form anti-antiterminator structures are underlined, those that form antiterminator structures are in italics, and those that form terminator structures are in boldface.
h Coding sequences are in upper case, and intergenic sequences are in lower case.
B. subtilis genes known or proposed to be regulated by anti-antiterminator, antiterminator, and terminator sequences and structures
a The homologous E. coli gene is also known to be regulated by attenuation.
b The homologous gene of E. coli was found by our program to have an attenuator.
c Gene regulated by the T-box mechanism ( 5 , 6 ).
d Gene regulated by the S-box mechanism ( 7 ).
e Gene regulated by TRAP (trp RNA–binding attenuation protein ( 19 ).
f Direction of transcription of the upstream gene. S, same direction, D, opposite direction.
g Sequences that form anti-antiterminator structures are underlined, those that form antiterminator structures are in italics, and those that form terminator structures are in boldface.
h Coding sequences are in upper case, and intergenic sequences are in lower case.
B. subtilis genes known or proposed to be regulated by anti-antiterminator, antiterminator, and terminator sequences and structures
a The homologous E. coli gene is also known to be regulated by attenuation.
b The homologous gene of E. coli was found by our program to have an attenuator.
c Gene regulated by the T-box mechanism ( 5 , 6 ).
d Gene regulated by the S-box mechanism ( 7 ).
e Gene regulated by TRAP (trp RNA–binding attenuation protein ( 19 ).
f Direction of transcription of the upstream gene. S, same direction, D, opposite direction.
g Sequences that form anti-antiterminator structures are underlined, those that form antiterminator structures are in italics, and those that form terminator structures are in boldface.
h Coding sequences are in upper case, and intergenic sequences are in lower case.
B. subtilis genes known or proposed to be regulated by anti-antiterminator, antiterminator, and terminator sequences and structures
a The homologous E. coli gene is also known to be regulated by attenuation.
b The homologous gene of E. coli was found by our program to have an attenuator.
c Gene regulated by the T-box mechanism ( 5 , 6 ).
d Gene regulated by the S-box mechanism ( 7 ).
e Gene regulated by TRAP (trp RNA–binding attenuation protein ( 19 ).
f Direction of transcription of the upstream gene. S, same direction, D, opposite direction.
g Sequences that form anti-antiterminator structures are underlined, those that form antiterminator structures are in italics, and those that form terminator structures are in boldface.
h Coding sequences are in upper case, and intergenic sequences are in lower case.
B. subtilis genes known or proposed to be regulated by anti-antiterminator, antiterminator, and terminator sequences and structures
a The homologous E. coli gene is also known to be regulated by attenuation.
b The homologous gene of E. coli was found by our program to have an attenuator.
c Gene regulated by the T-box mechanism ( 5 , 6 ).
d Gene regulated by the S-box mechanism ( 7 ).
e Gene regulated by TRAP (trp RNA–binding attenuation protein ( 19 ).
f Direction of transcription of the upstream gene. S, same direction, D, opposite direction.
g Sequences that form anti-antiterminator structures are underlined, those that form antiterminator structures are in italics, and those that form terminator structures are in boldface.
h Coding sequences are in upper case, and intergenic sequences are in lower case.
B. subtilis genes known or proposed to be regulated by anti-antiterminator, antiterminator, and terminator sequences and structures
a The homologous E. coli gene is also known to be regulated by attenuation.
b The homologous gene of E. coli was found by our program to have an attenuator.
c Gene regulated by the T-box mechanism ( 5 , 6 ).
d Gene regulated by the S-box mechanism ( 7 ).
e Gene regulated by TRAP (trp RNA–binding attenuation protein ( 19 ).
f Direction of transcription of the upstream gene. S, same direction, D, opposite direction.
g Sequences that form anti-antiterminator structures are underlined, those that form antiterminator structures are in italics, and those that form terminator structures are in boldface.
h Coding sequences are in upper case, and intergenic sequences are in lower case.
B. subtilis genes known or proposed to be regulated by anti-antiterminator, antiterminator, and terminator sequences and structures
a The homologous E. coli gene is also known to be regulated by attenuation.
b The homologous gene of E. coli was found by our program to have an attenuator.
c Gene regulated by the T-box mechanism ( 5 , 6 ).
d Gene regulated by the S-box mechanism ( 7 ).
e Gene regulated by TRAP (trp RNA–binding attenuation protein ( 19 ).
f Direction of transcription of the upstream gene. S, same direction, D, opposite direction.
g Sequences that form anti-antiterminator structures are underlined, those that form antiterminator structures are in italics, and those that form terminator structures are in boldface.
h Coding sequences are in upper case, and intergenic sequences are in lower case.
B. subtilis genes known or proposed to be regulated by anti-antiterminator, antiterminator, and terminator sequences and structures
a The homologous E. coli gene is also known to be regulated by attenuation.
b The homologous gene of E. coli was found by our program to have an attenuator.
c Gene regulated by the T-box mechanism ( 5 , 6 ).
d Gene regulated by the S-box mechanism ( 7 ).
e Gene regulated by TRAP (trp RNA–binding attenuation protein ( 19 ).
f Direction of transcription of the upstream gene. S, same direction, D, opposite direction.
g Sequences that form anti-antiterminator structures are underlined, those that form antiterminator structures are in italics, and those that form terminator structures are in boldface.
h Coding sequences are in upper case, and intergenic sequences are in lower case.
Homologous genes of B. subtilis and E. coli both of which appear to be regulated by transcription attenuation
a E. coli lysC is not regulated by attenuation ( 2 ), but its paralog, thrA, is regulated by antiterminator-terminator structures.
b Length of the fragments with maximum similarity in the alignment of their corresponding protein sequences.
c Percentage of identical residues common to the aligned protein sequences.
d Percentage of biochemically equivalent residues common to the aligned protein sequences.
Homologous genes of B. subtilis and E. coli both of which appear to be regulated by transcription attenuation
a E. coli lysC is not regulated by attenuation ( 2 ), but its paralog, thrA, is regulated by antiterminator-terminator structures.
b Length of the fragments with maximum similarity in the alignment of their corresponding protein sequences.
c Percentage of identical residues common to the aligned protein sequences.
d Percentage of biochemically equivalent residues common to the aligned protein sequences.