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Drosophila Kismet regulates histone H3 lysine 27 methylation and early elongation by RNA polymerase II.

Srinivasan S, Dorighi KM, Tamkun JW - PLoS Genet. (2008)

Bottom Line: Although we observed significant overlap between the distributions of KIS-L, ASH1, and TRX on polytene chromosomes, KIS-L did not bind methylated histone tails in vitro, and loss of TRX or ASH1 function did not alter the association of KIS-L with chromatin.By contrast, loss of kis function led to a dramatic reduction in the levels of TRX and ASH1 associated with chromatin and was accompanied by increased histone H3 lysine 27 methylation-a modification required for Polycomb group repression.Our findings suggest that KIS-L promotes early elongation and counteracts Polycomb group repression by recruiting the ASH1 and TRX histone methyltransferases to chromatin.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular, Cell, and Developmental Biology, University of California Santa Cruz, Santa Cruz, CA, USA.

ABSTRACT
Polycomb and trithorax group proteins regulate cellular pluripotency and differentiation by maintaining hereditable states of transcription. Many Polycomb and trithorax group proteins have been implicated in the covalent modification or remodeling of chromatin, but how they interact with each other and the general transcription machinery to regulate transcription is not well understood. The trithorax group protein Kismet-L (KIS-L) is a member of the CHD subfamily of chromatin-remodeling factors that plays a global role in transcription by RNA polymerase II (Pol II). Mutations in CHD7, the human counterpart of kis, are associated with CHARGE syndrome, a developmental disorder affecting multiple tissues and organs. To clarify how KIS-L activates gene expression and counteracts Polycomb group silencing, we characterized defects resulting from the loss of KIS-L function in Drosophila. These studies revealed that KIS-L acts downstream of P-TEFb recruitment to stimulate elongation by Pol II. The presence of two chromodomains in KIS-L suggested that its recruitment or function might be regulated by the methylation of histone H3 lysine 4 by the trithorax group proteins ASH1 and TRX. Although we observed significant overlap between the distributions of KIS-L, ASH1, and TRX on polytene chromosomes, KIS-L did not bind methylated histone tails in vitro, and loss of TRX or ASH1 function did not alter the association of KIS-L with chromatin. By contrast, loss of kis function led to a dramatic reduction in the levels of TRX and ASH1 associated with chromatin and was accompanied by increased histone H3 lysine 27 methylation-a modification required for Polycomb group repression. A similar increase in H3 lysine 27 methylation was observed in ash1 and trx mutant larvae. Our findings suggest that KIS-L promotes early elongation and counteracts Polycomb group repression by recruiting the ASH1 and TRX histone methyltransferases to chromatin.

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The association of KIS-L with chromatin is not altered in ash1 and trx mutants.A–D) The association of ASH1 (A, C, red) and KIS-L (B, D, green) on salivary gland polytene chromosomes of wild-type (A, B) and ash122/ash117 (C, D) larvae were detected by indirect immunofluorescence microscopy. E–H) The association of TRX (E, G, red) and KIS-L (F, H, green) on polytene chromosomes isolated from wild-type (E, F) and trx1 (G, H) larvae were detected by indirect immunofluorescence microscopy. Neither ASH1 nor TRX is required for the binding of KIS-L to polytene chromosomes.
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pgen-1000217-g005: The association of KIS-L with chromatin is not altered in ash1 and trx mutants.A–D) The association of ASH1 (A, C, red) and KIS-L (B, D, green) on salivary gland polytene chromosomes of wild-type (A, B) and ash122/ash117 (C, D) larvae were detected by indirect immunofluorescence microscopy. E–H) The association of TRX (E, G, red) and KIS-L (F, H, green) on polytene chromosomes isolated from wild-type (E, F) and trx1 (G, H) larvae were detected by indirect immunofluorescence microscopy. Neither ASH1 nor TRX is required for the binding of KIS-L to polytene chromosomes.

Mentions: The above results led us to question our hypothesis that ASH1 and TRX recruit KIS-L to chromatin by methylating H3K4 in the vicinity of promoters. To clarify this issue, we examined whether the loss of ASH1 or TRX function alters the association of KIS-L with salivary gland polytene chromosomes. Individuals trans-heterozygous for the hypomorphic ash122 and ash117 alleles survive until the third larval instar and display significantly reduced levels of ASH1 on polytene chromosomes (Figure 5A and C) [41]. No obvious changes in the level or distribution of KIS-L were observed in these mutants relative to wild-type (Figure 5B and D), indicating that ASH1 is not required for the association of KIS-L with chromatin. Similar results were obtained using a conditional trx allele, trx1. At 29°C, trx1 homozygotes survive until the third larval instar and display significantly reduced levels of TRX on polytene chromosomes (Figure 5E and G) [43]. We failed to detect obvious changes in the level or distribution of KIS-L on salivary gland chromosomes in trx1 mutants (Figure 5F and H). Thus, neither the ASH1 nor TRX histone methyltransferases are required for the association of KIS-L with chromatin in vivo.


Drosophila Kismet regulates histone H3 lysine 27 methylation and early elongation by RNA polymerase II.

Srinivasan S, Dorighi KM, Tamkun JW - PLoS Genet. (2008)

The association of KIS-L with chromatin is not altered in ash1 and trx mutants.A–D) The association of ASH1 (A, C, red) and KIS-L (B, D, green) on salivary gland polytene chromosomes of wild-type (A, B) and ash122/ash117 (C, D) larvae were detected by indirect immunofluorescence microscopy. E–H) The association of TRX (E, G, red) and KIS-L (F, H, green) on polytene chromosomes isolated from wild-type (E, F) and trx1 (G, H) larvae were detected by indirect immunofluorescence microscopy. Neither ASH1 nor TRX is required for the binding of KIS-L to polytene chromosomes.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC2563034&req=5

pgen-1000217-g005: The association of KIS-L with chromatin is not altered in ash1 and trx mutants.A–D) The association of ASH1 (A, C, red) and KIS-L (B, D, green) on salivary gland polytene chromosomes of wild-type (A, B) and ash122/ash117 (C, D) larvae were detected by indirect immunofluorescence microscopy. E–H) The association of TRX (E, G, red) and KIS-L (F, H, green) on polytene chromosomes isolated from wild-type (E, F) and trx1 (G, H) larvae were detected by indirect immunofluorescence microscopy. Neither ASH1 nor TRX is required for the binding of KIS-L to polytene chromosomes.
Mentions: The above results led us to question our hypothesis that ASH1 and TRX recruit KIS-L to chromatin by methylating H3K4 in the vicinity of promoters. To clarify this issue, we examined whether the loss of ASH1 or TRX function alters the association of KIS-L with salivary gland polytene chromosomes. Individuals trans-heterozygous for the hypomorphic ash122 and ash117 alleles survive until the third larval instar and display significantly reduced levels of ASH1 on polytene chromosomes (Figure 5A and C) [41]. No obvious changes in the level or distribution of KIS-L were observed in these mutants relative to wild-type (Figure 5B and D), indicating that ASH1 is not required for the association of KIS-L with chromatin. Similar results were obtained using a conditional trx allele, trx1. At 29°C, trx1 homozygotes survive until the third larval instar and display significantly reduced levels of TRX on polytene chromosomes (Figure 5E and G) [43]. We failed to detect obvious changes in the level or distribution of KIS-L on salivary gland chromosomes in trx1 mutants (Figure 5F and H). Thus, neither the ASH1 nor TRX histone methyltransferases are required for the association of KIS-L with chromatin in vivo.

Bottom Line: Although we observed significant overlap between the distributions of KIS-L, ASH1, and TRX on polytene chromosomes, KIS-L did not bind methylated histone tails in vitro, and loss of TRX or ASH1 function did not alter the association of KIS-L with chromatin.By contrast, loss of kis function led to a dramatic reduction in the levels of TRX and ASH1 associated with chromatin and was accompanied by increased histone H3 lysine 27 methylation-a modification required for Polycomb group repression.Our findings suggest that KIS-L promotes early elongation and counteracts Polycomb group repression by recruiting the ASH1 and TRX histone methyltransferases to chromatin.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular, Cell, and Developmental Biology, University of California Santa Cruz, Santa Cruz, CA, USA.

ABSTRACT
Polycomb and trithorax group proteins regulate cellular pluripotency and differentiation by maintaining hereditable states of transcription. Many Polycomb and trithorax group proteins have been implicated in the covalent modification or remodeling of chromatin, but how they interact with each other and the general transcription machinery to regulate transcription is not well understood. The trithorax group protein Kismet-L (KIS-L) is a member of the CHD subfamily of chromatin-remodeling factors that plays a global role in transcription by RNA polymerase II (Pol II). Mutations in CHD7, the human counterpart of kis, are associated with CHARGE syndrome, a developmental disorder affecting multiple tissues and organs. To clarify how KIS-L activates gene expression and counteracts Polycomb group silencing, we characterized defects resulting from the loss of KIS-L function in Drosophila. These studies revealed that KIS-L acts downstream of P-TEFb recruitment to stimulate elongation by Pol II. The presence of two chromodomains in KIS-L suggested that its recruitment or function might be regulated by the methylation of histone H3 lysine 4 by the trithorax group proteins ASH1 and TRX. Although we observed significant overlap between the distributions of KIS-L, ASH1, and TRX on polytene chromosomes, KIS-L did not bind methylated histone tails in vitro, and loss of TRX or ASH1 function did not alter the association of KIS-L with chromatin. By contrast, loss of kis function led to a dramatic reduction in the levels of TRX and ASH1 associated with chromatin and was accompanied by increased histone H3 lysine 27 methylation-a modification required for Polycomb group repression. A similar increase in H3 lysine 27 methylation was observed in ash1 and trx mutant larvae. Our findings suggest that KIS-L promotes early elongation and counteracts Polycomb group repression by recruiting the ASH1 and TRX histone methyltransferases to chromatin.

Show MeSH
Related in: MedlinePlus