Characterization of Cystathionine β‐Synthase TtCbs1 and Cysteine Synthase TtCsa1 Involved in Cysteine Biosynthesis in Tetrahymena thermophila

The thesis aims to identify and initiate functional characterization of the SWI/SNF and ISWI complexes in Tetrahymena thermophila. Through affinity purification of the conserved subunit Snf5 followed by mass spectrometry (AP-MS), I identified the first SWI/SNF complex in protists. One of the subunits I found is a small bromodomain containing protein named Ibd1. Through AP-MS of Ibd1 I found Ibd1 is versatile and interacts with several additional chromatin remodeling complexes. Bromodomains are known to have affinity for acetylated lysine residues within proteins such as histones. A peptide array experiment suggests that Ibd1 also has affinity for acetylated chromatin. Indirect immunofluorescence (IF) of Ibd1 hints at a role in transcription. My analysis of Tetrahymena Iswi1 shows expression during meiosis, vegetative growth and starvation. IF data shows its localization is consistent with Iswi1 function in mitosis/meiosis or maintenance of silent chromatin. AP-MS of ISW1 discovered several interacting proteins of unknown function.

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Molecular characterization of SerH3, a Tetrahymena thermophila gene encoding a temperature-regulated surface antigen

Molecular and Cellular Biology ◽

10.1128/mcb.10.11.6091-6096.1990 ◽

1990 ◽

Vol 10 (11) ◽

pp. 6091-6096

Author(s):

M M Tondravi ◽

R L Willis ◽

H D Love ◽

G A Bannon

Keyword(s):

Surface Antigen ◽

Dna Sequences ◽

Tetrahymena Thermophila ◽

Transcription Unit ◽

Genomic Fragment ◽

Gene Encoding ◽

Tryptophan Residues ◽

Functional Copy ◽

Regular Patterns

The DNA sequences of a cDNA clone and the macronuclear genomic fragment corresponding to the functional copy of the SerH3 surface antigen gene of Tetrahymena thermophila were determined. Primer extension and nuclease protection assays show that the SerH3 transcription unit is 1,425 nucleotides long and contains no introns. The predicted polypeptide encoded by the SerH3 gene has a molecular mass of 44,415 daltons; one-third of its 439 residues are either cysteine, serine, or threonine. The central half of the polypeptide consists of three homologous domains in tandem array; within these domains, the cysteine, proline, and tryptophan residues occur in highly regular patterns.

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Selection and characterization of a glucokinase-deficient mutant of Tetrahymena thermophila.

Molecular and Cellular Biology ◽

10.1128/mcb.2.4.378 ◽

1982 ◽

Vol 2 (4) ◽

pp. 378-385 ◽

Cited By ~ 2

Author(s):

C T Roberts ◽

J E Lavine ◽

D E Morse

Keyword(s):

Gene Expression ◽

Mutant Allele ◽

Tetrahymena Thermophila ◽

Molecular Defect ◽

Deficient Mutant ◽

Cellular Processes ◽

Inhibition Of Growth ◽

Selection Of ◽

Enzymatic Defect

We have isolated a mutant of Tetrahymena thermophila that is resistant to inhibition of growth by the glucose analog 2-deoxyglucose. The mutant exhibits a deficiency in a cytoplasmic glucokinase. This enzymatic defect and the attendant inability to convert 2-deoxyglucose to toxic phosphorylated derivatives is apparently the sole basis for the mutant phenotype since transport of glucose and 2-deoxyglucose is unimpaired; there is no elevation of glucose-6-phosphatase activity, which could decrease the level of toxic 2-deoxyglucose metabolites. Genetic analyses have shown that the mutant allele is recessive and inherited as a single Mendelian mutation. The glucokinase-deficient strain described here is useful for the selection of other mutants in this organism and for the investigation of various cellular processes initiated or modulated by glucose and its analogs. We have exploited the molecular defect in this strain to investigate the initial steps in the cyclic AMP-mediated repression of galactokinase gene expression which is caused by glucose.

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