MLL1 Promotes IL-7 Responsiveness and Survival during B Cell Differentiation.

B lymphocyte differentiation is an exquisitely regulated homeostatic process resulting in continuous production of appropriately selected B cells. Relatively small changes in gene expression can result in deregulation of this process, leading to acute lymphocytic leukemia (ALL), immune deficiency, or autoimmunity. Translocation of MLL1 (KMT2A) often results in a pro-B cell ALL, but little is known about its role in normal B cell differentiation. Using a Rag1-cre mouse knock-in to selectively delete Mll1 in developing lymphocytes, we show that B cell, but not T cell, homeostasis depends on MLL1. Mll1-/- B progenitors fail to differentiate efficiently through the pro- to pre-B cell transition, resulting in a persistent reduction in B cell populations. Cells inefficiently transit the pre-BCR checkpoint, despite normal to higher levels of pre-BCR components, and rearranged IgH expression fails to rescue this differentiation block. Instead of IgH-rearrangement defects, we find that Mll1-/- pre-B cells exhibit attenuated RAS/MAPK signaling downstream of the pre-BCR, which results in reduced survival in physiologic levels of IL-7. Genome-wide expression data illustrate that MLL1 is connected to B cell differentiation and IL-7-dependent survival through a complex transcriptional network. Overall, our data demonstrate that wild-type MLL1 is a regulator of pre-BCR signaling and B cell differentiation and further suggest that targeting its function in pro-B cell ALL may be more broadly effective than previously anticipated.

An MLL-dependent network sustains hematopoiesis.

The histone methyltransferase Mixed Lineage Leukemia (MLL) is essential to maintain hematopoietic stem cells and is a leukemia protooncogene. Although clustered homeobox genes are well-characterized targets of MLL and MLL fusion oncoproteins, the range of Mll-regulated genes in normal hematopoietic cells remains unknown. Here, we identify and characterize part of the Mll-dependent transcriptional network in hematopoietic stem cells with an integrated approach by using conditional loss-of-function models, genomewide expression analyses, chromatin immunoprecipitation, and functional rescue assays. The Mll-dependent transcriptional network extends well beyond the previously appreciated Hox targets, is comprised of many characterized regulators of self-renewal, and contains target genes that are both dependent and independent of the MLL cofactor, Menin. Interestingly, PR-domain containing 16 emerged as a target gene that is uniquely effective at partially rescuing Mll-deficient hematopoietic stem and progenitor cells. This work highlights the tissue-specific nature of regulatory networks under the control of MLL/Trithorax family members and provides insight into the distinctions between the participation of MLL in normal hematopoiesis and in leukemia.

Efficacy of the Atraumatic Physics Forceps Over Conventional Extraction Forceps in Extraction of Tooth-Does it Offer an Alternative in All Types of Extraction or Only can be Used in Few Selected Types of Extraction: A Comparative Study.

We conducted this study to evaluate the efficacy of physics forceps in dental extractions as compared with conventional forceps and as well as to evaluate its atraumatic point of view. Materials and Methods: A total of 200 healthy adult patients were included in the study who reported for dental extractions, divided into two groups each containing 100 patients. The patients were randomly allocated into either group by lottery method. Results: Operating time in the physics forceps group was found to be significantly reduced as well as lesser incidence of dry socket was noted which was statistically significant on the 2nd day postoperatively. However, incidence of root fracture, buccal plate fracture, and gingival lacerations were not found to be statistically significant but their occurrence was less in the physics forceps group as compared with the conventional forceps group. Conclusion: To conclude, we can say that extractions done by physics forceps results in significant reduction in unnecessary trauma as well as stress to both the operator and patient. The forceps is an atraumatic extraction system which is a boon for the profession, thereby reducing ridge deformities and preparing the patients for future prosthesis.

HOXA5 Expression Is Elevated in Breast Cancer and Is Transcriptionally Regulated by Estradiol.

HOXA5 is a homeobox-containing gene associated with the development of the lung, gastrointestinal tract, and vertebrae. Here, we investigate potential roles and the gene regulatory mechanism in HOXA5 in breast cancer cells. Our studies demonstrate that HOXA5 expression is elevated in breast cancer tissues and in estrogen receptor (ER)-positive breast cancer cells. HOXA5 expression is critical for breast cancer cell viability. Biochemical studies show that estradiol (E2) regulates HOXA5 gene expression in cultured breast cancer cells in vitro. HOXA5 expression is also upregulated in vivo in the mammary tissues of ovariectomized female rats. E2-induced HOXA5 expression is coordinated by ERs. Knockdown of either ERα or ERß downregulated E2-induced HOXA5 expression. Additionally, ER co-regulators, including CBP/p300 (histone acetylases) and MLL-histone methylases (MLL2, MLL3), histone acetylation-, and H3K4 trimethylation levels are enriched at the HOXA5 promoter in present E2. In summary, our studies demonstrate that HOXA5 is overexpressed in breast cancer and is transcriptionally regulated via estradiol in breast cancer cells.

Human CpG binding protein interacts with MLL1, MLL2 and hSet1 and regulates Hox gene expression.

Human encodes several histone H3-Lysine 4 (H3K4) specific methyl-transferases (HMTs) such as MLL1 (mixed lineage leukemia 1), MLL2, MLL3, hSet1 etc, that play critical roles in gene expression. These HMTs are present as distinct multi-protein complexes with several proteins in common. Herein, we have affinity purified and characterized human CpG binding protein (CGBP) and its interacting proteins from human cells. We demonstrated that CGBP is co-purified with three H3K4 specific HMTs MLL1, MLL2, and hSet1. We also performed independent immuno-precipitation of MLL1, MLL2 and hSet1 complexes from human cell and demonstrated that each of these complexes contains CGBP. In addition, CGBP is co-localized with MLL1, MLL2 and hSet1 in vivo and binds to the promoter of MLL target gene HoxA7. Antisense mediated knock down of CGBP diminished the recruitment of MLL1 and down regulated levels of H3K4 trimethylation in HoxA7 promoter affecting its expression. These results demonstrated that CGBP interacts with MLL1, MLL2 as well as hSet1 HMTs and plays critical roles in regulations of MLL target genes.

The histone methyltransferase activity of MLL1 is dispensable for hematopoiesis and leukemogenesis.

Despite correlations between histone methyltransferase (HMT) activity and gene regulation, direct evidence that HMT activity is responsible for gene activation is sparse. We address the role of the HMT activity for MLL1, a histone H3 lysine 4 (H3K4) methyltransferase critical for maintaining hematopoietic stem cells (HSCs). Here, we show that the SET domain, and thus HMT activity of MLL1, is dispensable for maintaining HSCs and supporting leukemogenesis driven by the MLL-AF9 fusion oncoprotein. Upon Mll1 deletion, histone H4 lysine 16 (H4K16) acetylation is selectively depleted at MLL1 target genes in conjunction with reduced transcription. Surprisingly, inhibition of SIRT1 is sufficient to prevent the loss of H4K16 acetylation and the reduction in MLL1 target gene expression. Thus, recruited MOF activity, and not the intrinsic HMT activity of MLL1, is central for the maintenance of HSC target genes. In addition, this work reveals a role for SIRT1 in opposing MLL1 function.

Dynamic association of MLL1, H3K4 trimethylation with chromatin and Hox gene expression during the cell cycle.

Mixed lineage leukemias (MLLs) are histone H3 at lysine 4 (H3K4)-specific methylases that play a critical role in regulating gene expression in humans. As chromatin condensation, relaxation and differential gene expression are keys to correct cell cycle progression, we analyzed the dynamic association of MLL and H3K4 trimethylation at different stages of the cell cycle. Interestingly, MLL1, which is normally associated with transcriptionally active chromatins (G1 phase), dissociates from condensed mitotic chromatin and returns at the end of telophase when the nucleus starts to relax. In contrast, H3K4 trimethylation mark, which is also normally associated with euchromatins (in G1), remains associated, even with condensed chromatin, throughout the cell cycle. The global levels of MLL1 and H3K4 trimethylation are not affected during the cell cycle, and H3Ser28 phosphorylation is only observed during mitosis. Interestingly, MLL target homeobox-containing (Hox) genes (HoxA5, HoxA7 and HoxA10) are differentially expressed during the cell cycle, and the recruitment of MLL1 and H3K4 trimethylation levels are modulated in the promoter of these Hox genes as a function of their expression. In addition, down-regulation of MLL1 results in cell cycle arrest at the G2/M phase. The fluctuation of H3K4 trimethylation marks at specific promoters, but not at the global level, indicates that H3K4 trimethylation marks that are present in the G1 phase may not be the same as the marks in other phases of the cell cycle; rather, old marks are removed and new marks are introduced. In conclusion, our studies demonstrate that MLL1 and H3K4 methylation have distinct dynamics during the cell cycle and play critical roles in the differential expression of Hox genes associated with cell cycle regulation.

MLL histone methylases in gene expression, hormone signaling and cell cycle.

Histone methyl-transferases (HMTs) are key enzymes that post-translationally methylate nuclear histone proteins and play critical roles in gene expression, epigenetic regulation and diseases in eukaryotic organisms. Mixed lineage leukemias (MLLs) are human HMTs that specifically methylate histone H3 at lyisine-4 and regulate gene activation. MLLs are also well known to be rearranged often in acute myeloid and lymphoid leukemias. Human encodes several MLLs that have similar enzymatic activities but diverse functions. Herein, we have reviewed the recent advances in understanding the diverse functions of MLL family of HMTs in gene regulation, hormone signaling and cell cycle regulation in human.