Proteolytic processing of micronuclear H3 and histone phosphorylation during conjugation in Tetrahymena thermophila

Allis, C. David; Wiggins, J C

doi:10.1016/0014-4827(84)90601-3

Cited by 39 publications

(18 citation statements)

References 19 publications

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“…Samples were loaded onto a 22% SDS-PAGE gel (60:0.4 acrylamide: bis-acrylamide) or for GLC7 experiments a 15% SDS-PAGE gel and blotted to polyvinylidene difluoride as described previously (29,30). Blots were blocked in 2-5% dry milk dissolved in 1ϫ TBST (0.15 M NaCl, 0.1 M Tris-HCl, pH7.5, 0.1%(w/v) Tween 20) for 2 h at room temperature or overnight at 4°C with agitation.…”

Section: Methodsmentioning

confidence: 99%

Site-specific Loss of Acetylation upon Phosphorylation of Histone H3

Edmondson¹,

Davie²,

Zhou³

et al. 2002

Journal of Biological Chemistry

102

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Post-translational modification of histones is a central aspect of gene regulation. Emerging data indicate that modification at one site can influence modification of a second site. As one example, histone H3 phosphorylation at serine 10 (Ser 10 ) facilitates acetylation of lysine 14 (Lys 14 ) by Gcn5 in vitro (1, 2). In vivo, phosphorylation of H3 precedes acetylation at certain promoters. Whether H3 phosphorylation globally affects acetylation, or whether it affects all acetylation sites in H3 equally, is not known. We have taken a genetic approach to this question by mutating Ser 10 in H3 to fix either a negative or a neutral charge at this position, followed by analysis of the acetylation states of the mutant histones using site-specific antibodies. Surprisingly, we find that conversion of Ser 10 to glutamate (S10E) or aspartate (S10D) causes almost complete loss of H3 acetylation at lysine 9 (Lys 9 ) in vivo. Acetylation of Lys 9 is also significantly reduced in cells bearing mutations in the Glc7 phosphatase that increase H3 phosphorylation levels. Mutation of Ser 10 in H3 and the concomitant loss of Lys 9 acetylation has minimal effects on expression of a Gcn5-dependent reporter gene. However, synergistic growth defects are observed upon loss of GCN5 in cells bearing H3 Ser 10 mutations that are reminiscent of delays in G 2 /M progression caused by combined loss of GCN5 and acetylation site mutations. Together these results demonstrate that H3 phosphorylation directly causes sitespecific and opposite changes in acetylation levels of two residues within this histone, Lys 9 and Lys 14 , and they highlight the importance of these histone modifications to normal cell functions.

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Section: Methodsmentioning

confidence: 99%

Site-specific Loss of Acetylation upon Phosphorylation of Histone H3

Edmondson¹,

Davie²,

Zhou³

et al. 2002

Journal of Biological Chemistry

102

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“…Throughout this period, newly synthesized histones were deacetylated by an unknown micronuclear HDAC (2,8). Following deacetylation, micronuclear chromatin is thought to mature through evolutionarily conserved pathways, but one unique feature is the proteolytic processing of a fraction of new histone H3 molecules into a truncated form (3,5,6). Production of this form is related to mitotic chromosome condensation and segregation in the micronucleus (54).…”

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confidence: 99%

“…In micronuclei, histone H3 (called H3 s ) is proteolytically cleaved to form a faster-migrating form, called H3 f , which is missing 6 amino acids from the extreme amino terminus. H3 f is the only form of histone H3 in Tetrahymena that is phosphorylated on Ser10, a mitotic modification necessary for chromosome condensation and segregation (4,6,54). In thd2⌬ cells, the elongated micronuclear phenotype, reminiscent of either decondensed chromatin or mitotic micronuclei in anaphase, prompted us to examine H3 processing and related phosphorylation on Ser10.…”

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confidence: 99%

A Class II Histone Deacetylase Acts on Newly Synthesized Histones in Tetrahymena

et al. 2008

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Newly synthesized histones are acetylated prior to their deposition into nucleosomes. Following nucleosome formation and positioning, they are rapidly deacetylated, an event that coincides with further maturation of the chromatin fiber. The histone deacetylases (HDACs) used for histone deposition and de novo chromatin formation are poorly understood. In the ciliate Tetrahymena thermophila, transcription-related deacetylation in the macronucleus is physically separated from deposition-related deacetylation in the micronucleus. This feature was utilized to identify an HDAC named Thd2, a class II HDAC that acts on newly synthesized histones to remove deposition-related acetyl moieties. The THD2 transcript is alternatively spliced, and the major form contains a putative inositol polyphosphate kinase (IPK) domain similar to Ipk2, an enzyme that promotes chromatin remodeling by SWI/SNF remodeling complexes. Cells lacking Thd2, which retain deposition-related acetyl moieties on new histones, exhibit chromatin and cytological phenotypes indicative of a role for Thd2 in chromatin maturation, including the proteolytic processing of histone H3.

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“…The chromatin of two formerly completely silent micronuclei is committed to transcriptional activity whereas the other two micronuclei remain inactive and develop to maintain the genetic continuity of the organism including the ability to undergo meiosis in future conjugations. These processes are accompanied by a series of events such as selective deposition ofhistone variants (39) and proteolytic processing of H3 and H1 like histones (3,4,6,12). Furthermore, new macronuclei undergo large scale genome rearrangements accompanied by elimination of micronuclear sequences, gene amplification and formation of new telomeres (for a review see reference 4).…”

Section: Discussionmentioning

confidence: 99%

Histone acetylation in conjugating Tetrahymena thermophila [published erratum appears in J Cell Biol 1989 Dec;109(6 Pt 1):3214-7]

Pfeffer¹

1989

The Journal of Cell Biology

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Abstract. We have monitored histone acetylation during conjugation of the ciliated protozoan Tetrahymena thermophila using antibodies against the tetraacetylated form of H4 histone (Pfeffer, U., N. Ferrari, and G. Vidali. 1986. J. Biol. Chem. 261:2496-2498. During meiosis, the three prezygotic divisions, fertilization, and the first postzygotic division, micronuclei, do not contain highly acetylated forms of H4 histone. However, after the second postzygotic division, when anteriorly located micronuclei begin to develop into new macronuclei, they are strongly stained by the anti-tetraacetylated H4 histone antibody. In the old macronucleus, histones are actively deacetylated when it has ceased to transcribe but before it is eliminated. Histone acetylation processes analyzed here appear to be correlated to the commitment to transcription rather than to the transcription process itself. This is in good correlation with evidence we have obtained in chick erythrocyte nuclei during reactivation upon fusion with mammalian cells (Pfeffer, U., N. Ferrari, E Tosetti, and G. Vidali. 1988. Exp. Cell Res. 178:25-30). Furthermore, it becomes clear from our data that histone acetylation occurs in close correlation to the position of nuclei within the cytoplasm of T. thermophila.Mechanisms that control differential histone acetylation and deacetylation are discussed.S INCE the original observation, histone acetylation has been correlated to transcription and considerable evidence has accumulated indicating that this reversible postsynthetic modification of lysine residues in the aminoterminal portion of core histones plays a key role in the regulation of gene expression (for a review see reference 2).Different methods have been developed to fractionate chromatin depending on its transcriptional activity and accordingly, it has been found that fractions containing actively transcribed genes were enriched in highly acetylated histones (1,17,33). Recent analyses using anti-acetyl-lysine antibodies have established that a given gene is associated with highly acetylated histones when it is transcribed (23), although it cannot be ruled out that genes competent to be transcribed, but inactive at the moment of analysis, may also contain acetylated histones.Nucleosomes undergo reversible conformational changes during transcriptional activation (10, 32), histones are absent from promoter sequences of active genes (25), nucleosomes inhibit initiation of transcription but do not inhibit elongation of initiated transcripts (27, 28) and at least H4 histone is retained on a highly transcribed gene (35). The contact points between nucleosomal DNA and H4 histone have been analyzed and a highly basic cluster in the amino-terminal portion has been shown to be involved in sharp bending of the DNA double helix (18). Interestingly, this cluster encompasses lysine at position 16 which is the first to be acetylated in vivo in calf thymus (15) and, moreover, this region is in close contact to a site 55-65 bp from the end of the nucleosomal DNA, which ...

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Proteolytic processing of micronuclear H3 and histone phosphorylation during conjugation in Tetrahymena thermophila

Cited by 39 publications

References 19 publications

Site-specific Loss of Acetylation upon Phosphorylation of Histone H3

Site-specific Loss of Acetylation upon Phosphorylation of Histone H3

A Class II Histone Deacetylase Acts on Newly Synthesized Histones in Tetrahymena

Histone acetylation in conjugating Tetrahymena thermophila [published erratum appears in J Cell Biol 1989 Dec;109(6 Pt 1):3214-7]

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