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Inhibition of class I HDACs abrogates the dominant effect of MLL-AF4 by activation of wild-type MLL.

Ahmad K, Katryniok C, Scholz B, Merkens J, Löscher D, Marschalek R, Steinhilber D - Oncogenesis (2014)

Bottom Line: Transcriptional initiation was associated with an increase in histone H3 lysine 4 trimethylation in a TSA-inducible manner.Therefore, we investigated the effects of the MLL (mixed lineage leukemia) protein and its derivatives, MLL-AF4 and AF4-MLL, respectively.Surprisingly, a constitutive activation of ALOX5 by MLL-AF4 was inhibited by class I HDAC inhibitors, by relieving inhibitory functions deriving from MLL.Conversely, a knockdown of MLL increased the effects mediated by MLL-AF4.

View Article: PubMed Central - PubMed

Affiliation: Institute of Pharmaceutical Chemistry/ZAFES, Goethe University Frankfurt, Frankfurt, Germany.

ABSTRACT
The ALOX5 gene encodes 5-lipoxygenase (5-LO), a key enzyme of inflammatory reactions, which is transcriptionally activated by trichostatin A (TSA). Physiologically, 5-LO expression is induced by calcitriol and/or transforming growth factor-β. Regulation of 5-LO mRNA involves promoter activation and elongation control within the 3'-portion of the ALOX5 gene. Here we focused on the ALOX5 promoter region. Transcriptional initiation was associated with an increase in histone H3 lysine 4 trimethylation in a TSA-inducible manner. Therefore, we investigated the effects of the MLL (mixed lineage leukemia) protein and its derivatives, MLL-AF4 and AF4-MLL, respectively. MLL-AF4 was able to enhance ALOX5 promoter activity by 47-fold, which was further stimulated when either vitamin D receptor and retinoid X receptor or SMAD3/SMAD4 were co-transfected. In addition, we investigated several histone deacetylase inhibitors (HDACi) in combination with gene knockdown experiments (HDAC1-3, MLL). We were able to demonstrate that a combined inhibition of HDAC1-3 induces ALOX5 promoter activity in an MLL-dependent manner. Surprisingly, a constitutive activation of ALOX5 by MLL-AF4 was inhibited by class I HDAC inhibitors, by relieving inhibitory functions deriving from MLL.Conversely, a knockdown of MLL increased the effects mediated by MLL-AF4. Thus, HDACi treatment seems to switch 'inactive MLL' into 'active MLL' and overwrites the dominant functions deriving from MLL-AF4.

No MeSH data available.


Related in: MedlinePlus

Proposed model for the regulation of 5-LO promoter activity by MLL and MLL-AF4. Top: the MLL protein complex can act as an activator or a repressor of transcription, depending on the incorporation of CYP33. This protein modulates the PHD3 domain of MLL in a way that the domain either reads H3K4 signatures or enables the association of MLL with a repressor complex (BMI-1, HDACs and so on, respectively). The constitutively active MLL-AF4 fusion protein (which lacks the regulatory PHD3 domain) overrides the endogenous MLL protein leading to aberrant transcriptional activity. Bottom: in the presence of class I HDACi that inhibit HDACs1–3, the MLL complex loses its inhibitory regulatory function, which in turn allows the MLL complex to compete with MLL-AF4 and to diminish the (oncogenic) functions of the MLL-AF4 oncoprotein on transcription.
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fig9: Proposed model for the regulation of 5-LO promoter activity by MLL and MLL-AF4. Top: the MLL protein complex can act as an activator or a repressor of transcription, depending on the incorporation of CYP33. This protein modulates the PHD3 domain of MLL in a way that the domain either reads H3K4 signatures or enables the association of MLL with a repressor complex (BMI-1, HDACs and so on, respectively). The constitutively active MLL-AF4 fusion protein (which lacks the regulatory PHD3 domain) overrides the endogenous MLL protein leading to aberrant transcriptional activity. Bottom: in the presence of class I HDACi that inhibit HDACs1–3, the MLL complex loses its inhibitory regulatory function, which in turn allows the MLL complex to compete with MLL-AF4 and to diminish the (oncogenic) functions of the MLL-AF4 oncoprotein on transcription.

Mentions: Interestingly, the MLL-AF4 fusion protein still retains the MT domain, but lacks the PHD domain. According to our data, MLL-AF4 acts similar to a constitutively active version of the MLL protein on the tested ALOX5 promoter. Addition of different HDACi that specifically block HDAC1–3 activities allows efficient activation of the reporter construct by endogenous MLL, while binding to and activation of the ALOX5 promoter by MLL-AF4 is concomitantly reduced. This mode of action is displayed in Figure 9, where class I HDACs (presumably HDAC1 and 2) seem to act on the activator/repressor switch of MLL, which significantly reduce binding of MLL-AF4 to the ALOX5 promoter. Interestingly, the presence of MLL-AF4 seems to dramatically enhance the binding of MLL. This may indicate an additional and novel function of MLL-AF4, which could be ‘chromatin opening', for example, by recruiting the SWI/SNF complex. This needs to be investigated in future experiments.


Inhibition of class I HDACs abrogates the dominant effect of MLL-AF4 by activation of wild-type MLL.

Ahmad K, Katryniok C, Scholz B, Merkens J, Löscher D, Marschalek R, Steinhilber D - Oncogenesis (2014)

Proposed model for the regulation of 5-LO promoter activity by MLL and MLL-AF4. Top: the MLL protein complex can act as an activator or a repressor of transcription, depending on the incorporation of CYP33. This protein modulates the PHD3 domain of MLL in a way that the domain either reads H3K4 signatures or enables the association of MLL with a repressor complex (BMI-1, HDACs and so on, respectively). The constitutively active MLL-AF4 fusion protein (which lacks the regulatory PHD3 domain) overrides the endogenous MLL protein leading to aberrant transcriptional activity. Bottom: in the presence of class I HDACi that inhibit HDACs1–3, the MLL complex loses its inhibitory regulatory function, which in turn allows the MLL complex to compete with MLL-AF4 and to diminish the (oncogenic) functions of the MLL-AF4 oncoprotein on transcription.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4259963&req=5

fig9: Proposed model for the regulation of 5-LO promoter activity by MLL and MLL-AF4. Top: the MLL protein complex can act as an activator or a repressor of transcription, depending on the incorporation of CYP33. This protein modulates the PHD3 domain of MLL in a way that the domain either reads H3K4 signatures or enables the association of MLL with a repressor complex (BMI-1, HDACs and so on, respectively). The constitutively active MLL-AF4 fusion protein (which lacks the regulatory PHD3 domain) overrides the endogenous MLL protein leading to aberrant transcriptional activity. Bottom: in the presence of class I HDACi that inhibit HDACs1–3, the MLL complex loses its inhibitory regulatory function, which in turn allows the MLL complex to compete with MLL-AF4 and to diminish the (oncogenic) functions of the MLL-AF4 oncoprotein on transcription.
Mentions: Interestingly, the MLL-AF4 fusion protein still retains the MT domain, but lacks the PHD domain. According to our data, MLL-AF4 acts similar to a constitutively active version of the MLL protein on the tested ALOX5 promoter. Addition of different HDACi that specifically block HDAC1–3 activities allows efficient activation of the reporter construct by endogenous MLL, while binding to and activation of the ALOX5 promoter by MLL-AF4 is concomitantly reduced. This mode of action is displayed in Figure 9, where class I HDACs (presumably HDAC1 and 2) seem to act on the activator/repressor switch of MLL, which significantly reduce binding of MLL-AF4 to the ALOX5 promoter. Interestingly, the presence of MLL-AF4 seems to dramatically enhance the binding of MLL. This may indicate an additional and novel function of MLL-AF4, which could be ‘chromatin opening', for example, by recruiting the SWI/SNF complex. This needs to be investigated in future experiments.

Bottom Line: Transcriptional initiation was associated with an increase in histone H3 lysine 4 trimethylation in a TSA-inducible manner.Therefore, we investigated the effects of the MLL (mixed lineage leukemia) protein and its derivatives, MLL-AF4 and AF4-MLL, respectively.Surprisingly, a constitutive activation of ALOX5 by MLL-AF4 was inhibited by class I HDAC inhibitors, by relieving inhibitory functions deriving from MLL.Conversely, a knockdown of MLL increased the effects mediated by MLL-AF4.

View Article: PubMed Central - PubMed

Affiliation: Institute of Pharmaceutical Chemistry/ZAFES, Goethe University Frankfurt, Frankfurt, Germany.

ABSTRACT
The ALOX5 gene encodes 5-lipoxygenase (5-LO), a key enzyme of inflammatory reactions, which is transcriptionally activated by trichostatin A (TSA). Physiologically, 5-LO expression is induced by calcitriol and/or transforming growth factor-β. Regulation of 5-LO mRNA involves promoter activation and elongation control within the 3'-portion of the ALOX5 gene. Here we focused on the ALOX5 promoter region. Transcriptional initiation was associated with an increase in histone H3 lysine 4 trimethylation in a TSA-inducible manner. Therefore, we investigated the effects of the MLL (mixed lineage leukemia) protein and its derivatives, MLL-AF4 and AF4-MLL, respectively. MLL-AF4 was able to enhance ALOX5 promoter activity by 47-fold, which was further stimulated when either vitamin D receptor and retinoid X receptor or SMAD3/SMAD4 were co-transfected. In addition, we investigated several histone deacetylase inhibitors (HDACi) in combination with gene knockdown experiments (HDAC1-3, MLL). We were able to demonstrate that a combined inhibition of HDAC1-3 induces ALOX5 promoter activity in an MLL-dependent manner. Surprisingly, a constitutive activation of ALOX5 by MLL-AF4 was inhibited by class I HDAC inhibitors, by relieving inhibitory functions deriving from MLL.Conversely, a knockdown of MLL increased the effects mediated by MLL-AF4. Thus, HDACi treatment seems to switch 'inactive MLL' into 'active MLL' and overwrites the dominant functions deriving from MLL-AF4.

No MeSH data available.


Related in: MedlinePlus