Cloning of mtr, an amino acid transport gene of Neurospora crassa

Genome ◽  
1988 ◽  
Vol 30 (2) ◽  
pp. 198-203 ◽  
Author(s):  
W. Dorsey Stuart ◽  
Kenneth Koo ◽  
Steven J. Vollmer
Keyword(s):  
Amino Acids ◽  
Neurospora Crassa ◽  
Messenger Rna ◽  
Genomic Dna ◽  
Gene Sequence ◽  
Fragment Length Polymorphism ◽  
Aromatic Amino Acids ◽  
Cosmid Library ◽  
Coding Region ◽  
Rna Transcript

Translocation of neutral aliphatic and aromatic amino acids across the plasma membrane of the ascomycete Neurospora crassa requires a functional gene product of the mtr locus. Mutations at this locus are defective in transport of those amino acids. We have cloned the mtr+ gene of Neurospora crassa from an ordered cosmid library of genomic DNA and produced a preliminary restriction map of 2.9 kilobases of genomic DNA that encompasses the mtr coding region. We have confirmed that the cloned DNA regions contain the mtr gene sequence by restriction fragment length polymorphism mapping and have determined that the cloned sequence codes for a messenger RNA transcript of approximately 1200 nucleotides in length.Key words: transformation, methyltryptophane resistance, Neurospora.

scholarly journals Cloning and characterization of platelet factor 4 cDNA derived from a human erythroleukemic cell line

Blood ◽  
1987 ◽  
Vol 69 (1) ◽  
pp. 219-223 ◽  
Author(s):  
M Poncz ◽  
S Surrey ◽  
P LaRocco ◽  
MJ Weiss ◽  
EF Rappaport ◽  
...  
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Cell Line ◽  
Genomic Dna ◽  
Cdna Probe ◽  
Leader Sequence ◽  
Platelet Factor 4 ◽  
Coding Region ◽  
Platelet Factor ◽  
Amino Acid Homology

Abstract We report the isolation of a platelet factor 4 (PF4) cDNA clone from a lambda gt11 expression cDNA library which was derived from a human erythroleukemic (HEL) cell line. The sequence of the DNA insert includes the 3′-untranslated region, the entire amino acid coding region for the mature PF4 protein, and a 5′ region containing coding information for an additional 18 amino acids. In addition, supplemental genomic DNA sequencing shows that the full-length leader sequence is 30 amino acids long plus an initial methionine and codes for a hydrophobic signal-like sequence which is probably involved in transmembrane transport. A single species mRNA of approximately 800 nucleotides was detected on blots of HEL cell poly(A) + RNA using a labeled PF4 cDNA probe. The human PF4 leader sequence shares some DNA, but no amino acid, homology with the 15 amino acids at the N-terminus of mature bovine PF4, suggesting rapid divergence in this region of PF4 between these two species. Sequence comparison of the coding regions of mature PF4 and gamma IP-10, a protein induced in a variety of cells following treatment with gamma-interferon, shows a corrected divergence of 76%. The divergence of a common ancestor protein into PF4 and gamma IP-10 may have accompanied the development of sophisticated immune and coagulation systems in vertebrates. The availability of cDNA and genomic DNA information for these genes in other species will be useful in studying the evolution of the coagulation and immune systems.

scholarly journals Cloning and characterization of platelet factor 4 cDNA derived from a human erythroleukemic cell line

Blood ◽  
1987 ◽  
Vol 69 (1) ◽  
pp. 219-223 ◽  
Author(s):  
M Poncz ◽  
S Surrey ◽  
P LaRocco ◽  
MJ Weiss ◽  
EF Rappaport ◽  
...  
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Cell Line ◽  
Genomic Dna ◽  
Cdna Probe ◽  
Leader Sequence ◽  
Platelet Factor 4 ◽  
Coding Region ◽  
Platelet Factor ◽  
Amino Acid Homology

We report the isolation of a platelet factor 4 (PF4) cDNA clone from a lambda gt11 expression cDNA library which was derived from a human erythroleukemic (HEL) cell line. The sequence of the DNA insert includes the 3′-untranslated region, the entire amino acid coding region for the mature PF4 protein, and a 5′ region containing coding information for an additional 18 amino acids. In addition, supplemental genomic DNA sequencing shows that the full-length leader sequence is 30 amino acids long plus an initial methionine and codes for a hydrophobic signal-like sequence which is probably involved in transmembrane transport. A single species mRNA of approximately 800 nucleotides was detected on blots of HEL cell poly(A) + RNA using a labeled PF4 cDNA probe. The human PF4 leader sequence shares some DNA, but no amino acid, homology with the 15 amino acids at the N-terminus of mature bovine PF4, suggesting rapid divergence in this region of PF4 between these two species. Sequence comparison of the coding regions of mature PF4 and gamma IP-10, a protein induced in a variety of cells following treatment with gamma-interferon, shows a corrected divergence of 76%. The divergence of a common ancestor protein into PF4 and gamma IP-10 may have accompanied the development of sophisticated immune and coagulation systems in vertebrates. The availability of cDNA and genomic DNA information for these genes in other species will be useful in studying the evolution of the coagulation and immune systems.

scholarly journals Ornithine decarboxylase gene of Neurospora crassa: isolation, sequence, and polyamine-mediated regulation of its mRNA

1992 ◽  
Vol 12 (1) ◽  
pp. 347-359
Author(s):  
L J Williams ◽  
G R Barnett ◽  
J L Ristow ◽  
J Pitkin ◽  
M Perriere ◽  
...  
Keyword(s):  
Amino Acids ◽  
Neurospora Crassa ◽  
Ornithine Decarboxylase ◽  
Amino Acid Identity ◽  
Coding Region ◽  
Reading Frame ◽  
Translational Start ◽  
Rna Blots ◽  
Striking Contrast

Ornithine decarboxylase (ODC), which initiates the biosynthesis of the polyamines putrescine, spermidine, and spermine, is encoded by the spe-1 gene of the fungus Neurospora crassa. This gene and its cDNA have been cloned and sequenced. The gene has a single 70-nucleotide intron in the coding sequence. The cDNA, comprising the entire coding region, recognizes a single 2.4-kb mRNA in Northern (RNA) blots. The mRNA transcript, defined by S1 mapping, has an extremely long, 535-base leader without strong secondary-structure features or an upstream reading frame. The translational start of the protein is ambiguous: a Met-Val-Met sequence precedes the Pro known to be the N terminus of the ODC polypeptide. The polypeptide encoded by the N. crassa spe-1 gene (484 amino acids) has 46% amino acid identity with that of Saccharomyces cerevisiae (466 amino acids) and 42% with that of mouse (461 amino acids). Alignment of the longer N. crassa sequence with S. cerevisiae and mouse sequences creates gaps in different sites in the S. cerevisiae and mouse sequences, suggesting that N. crassa ODC is closer to an ancestral form of the enzyme than that of either yeast or mouse ODC. N. crassa ODC, which turns over rapidly in vivo in the presence of polyamines, has two PEST sequences, found in most ODCs and other proteins with rapid turnover. In striking contrast to other eucaryotic organisms, the variation in the rate of ODC synthesis in response to polyamines in N. crassa is largely correlated with proportional changes in the abundance of ODC mRNA. Spermidine is the main effector of repression, while putrescine has a weaker effect. However, putrescine accumulation appears to increase the amount of active ODC that is made from a given amount of ODC mRNA, possibly by improving its translatability. Conversely, prolonged starvation for both putrescine and spermidine leads to the differentially impaired translation of ODC mRNA.

scholarly journals Ornithine decarboxylase gene of Neurospora crassa: isolation, sequence, and polyamine-mediated regulation of its mRNA.

1992 ◽  
Vol 12 (1) ◽  
pp. 347-359 ◽  
Author(s):  
L J Williams ◽  
G R Barnett ◽  
J L Ristow ◽  
J Pitkin ◽  
M Perriere ◽  
...  
Keyword(s):  
Amino Acids ◽  
Neurospora Crassa ◽  
Ornithine Decarboxylase ◽  
Amino Acid Identity ◽  
Coding Region ◽  
Reading Frame ◽  
Translational Start ◽  
Rna Blots ◽  
Striking Contrast

Ornithine decarboxylase (ODC), which initiates the biosynthesis of the polyamines putrescine, spermidine, and spermine, is encoded by the spe-1 gene of the fungus Neurospora crassa. This gene and its cDNA have been cloned and sequenced. The gene has a single 70-nucleotide intron in the coding sequence. The cDNA, comprising the entire coding region, recognizes a single 2.4-kb mRNA in Northern (RNA) blots. The mRNA transcript, defined by S1 mapping, has an extremely long, 535-base leader without strong secondary-structure features or an upstream reading frame. The translational start of the protein is ambiguous: a Met-Val-Met sequence precedes the Pro known to be the N terminus of the ODC polypeptide. The polypeptide encoded by the N. crassa spe-1 gene (484 amino acids) has 46% amino acid identity with that of Saccharomyces cerevisiae (466 amino acids) and 42% with that of mouse (461 amino acids). Alignment of the longer N. crassa sequence with S. cerevisiae and mouse sequences creates gaps in different sites in the S. cerevisiae and mouse sequences, suggesting that N. crassa ODC is closer to an ancestral form of the enzyme than that of either yeast or mouse ODC. N. crassa ODC, which turns over rapidly in vivo in the presence of polyamines, has two PEST sequences, found in most ODCs and other proteins with rapid turnover. In striking contrast to other eucaryotic organisms, the variation in the rate of ODC synthesis in response to polyamines in N. crassa is largely correlated with proportional changes in the abundance of ODC mRNA. Spermidine is the main effector of repression, while putrescine has a weaker effect. However, putrescine accumulation appears to increase the amount of active ODC that is made from a given amount of ODC mRNA, possibly by improving its translatability. Conversely, prolonged starvation for both putrescine and spermidine leads to the differentially impaired translation of ODC mRNA.

Gene disruption by transformation in Neurospora crassa

1985 ◽  
Vol 5 (7) ◽  
pp. 1554-1559
Author(s):  
J V Paietta ◽  
G A Marzluf
Keyword(s):  
Neurospora Crassa ◽  
Plasmid Dna ◽  
Genomic Dna ◽  
Gene Disruption ◽  
Transformation Frequency ◽  
Coding Region ◽  
Multiple Integration ◽  
Southern Blots ◽  
Homologous Site ◽  
Hybridization Band

To establish conditions which might permit deliberate gene disruptions in Neurospora crassa, we studied transformation with linear DNA fragments. The transformation frequency observed was increased about twofold in comparison with that obtained with circular plasmid DNA. However, only a low proportion, approximately 10%, of the integration events occurred at the homologous site, whereas most integrations of transforming DNA took place in nonhomologous regions. It was also found that multiple integration events frequently occurred in individual transformants. A plasmid, designated pJP12, was constructed that contains the N. crassa am+ gene interrupted by insertion into its coding region of a DNA segment carrying a functional Neurospora qa-2+ gene. A fragment of Neurospora DNA that contains this am qa-2+ construction was obtained from plasmid pJP12 and used to transform an am+ qa-2 strain in an attempt to disrupt the resident am+ gene. After the initial qa-2+ transformants were converted to homokaryons by appropriate crosses, 10 independent transformants with an am mutant phenotype were found among 117 examined. Each of these qa-2+ am transformants showed the loss of a hybridization band in Southern blots of genomic DNA that corresponded to the normal am+ gene and the presence of a new hybridization band, consistent with an alteration in the am+ region.

scholarly journals Gene disruption by transformation in Neurospora crassa.

1985 ◽  
Vol 5 (7) ◽  
pp. 1554-1559 ◽  
Author(s):  
J V Paietta ◽  
G A Marzluf
Keyword(s):  
Neurospora Crassa ◽  
Plasmid Dna ◽  
Genomic Dna ◽  
Gene Disruption ◽  
Transformation Frequency ◽  
Coding Region ◽  
Multiple Integration ◽  
Southern Blots ◽  
Homologous Site ◽  
Hybridization Band

To establish conditions which might permit deliberate gene disruptions in Neurospora crassa, we studied transformation with linear DNA fragments. The transformation frequency observed was increased about twofold in comparison with that obtained with circular plasmid DNA. However, only a low proportion, approximately 10%, of the integration events occurred at the homologous site, whereas most integrations of transforming DNA took place in nonhomologous regions. It was also found that multiple integration events frequently occurred in individual transformants. A plasmid, designated pJP12, was constructed that contains the N. crassa am+ gene interrupted by insertion into its coding region of a DNA segment carrying a functional Neurospora qa-2+ gene. A fragment of Neurospora DNA that contains this am qa-2+ construction was obtained from plasmid pJP12 and used to transform an am+ qa-2 strain in an attempt to disrupt the resident am+ gene. After the initial qa-2+ transformants were converted to homokaryons by appropriate crosses, 10 independent transformants with an am mutant phenotype were found among 117 examined. Each of these qa-2+ am transformants showed the loss of a hybridization band in Southern blots of genomic DNA that corresponded to the normal am+ gene and the presence of a new hybridization band, consistent with an alteration in the am+ region.

The Efficient Preparation of Radiolabeled Aromatic Amino Acids via Cu-Mediated Radiofluorination using Ni-complexes

2019 ◽  
Author(s):  
A Craig ◽  
N Kolks ◽  
E Urusova ◽  
BD Zlatopolskiy ◽  
B Neumaier
Keyword(s):  
Amino Acids ◽  
Aromatic Amino Acids

Long term dietary intake of aromatic amino acids are associated with leptin gene expression in adipose tissues of non-diabetic adults

2018 ◽  
Author(s):  
Golaleh Asghari ◽  
Emad Yuzbashian ◽  
Maryam Zarkesh ◽  
Parvin Mirmiran ◽  
Mehdi Hedayati ◽  
...  
Keyword(s):  
Gene Expression ◽  
Amino Acids ◽  
Dietary Intake ◽  
Aromatic Amino Acids ◽  
Adipose Tissues
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