Klimczak, M. | Search bioRxiv

bioRxiv2021

Clustered PHD domains in mixed lineage leukaemia proteins are attracted by acetylation-rich active promoters and enhancers

Histone lysine-specfic methyltransferase 2 (KMT2A-D) proteins, alternatively called mixed lineage leukaemia (MLL1-4) proteins, mediate positive transcriptional memory. As the catalytic subunits of human COMPASS-like complexes, they methylate H3K4 at promoters and enhancers. KMT2A-D contain understudied highly conserved triplets and a quartet of plant homeodomains (PHDs). Here, we show that all clustered PHDs localise to the well-defined loci of H3K4me3 and H3 acetylation-rich active promoters and enhancers. Surprisingly, we observe little difference in binding pattern between PHDs from promoter-specific KMT2A-B and enhancer-specific KMT2C-D. Fusion of the KMT2A CXXC domain to the PHDs drastically enhances their preference for promoters over enhancers. Hence, the presence of CXXC domains in KMT2A-B, but not KMT2C-D, may explain the promoter/enhancer preferences of the full-length proteins. Importantly, targets of PHDs overlap with KMT2A targets and are enriched in genes involved in the cancer pathways. We also observe that PHDs of KMT2A-D are mutated in cancer, especially within conserved folding motifs (Cys4HisCys2Cys/His), which cause a domain loss-of-function. Taken together, our data suggests that PHDs of KMT2A-D guide the full-length proteins to active promoters and enhancers, and thus play a role in positive transcriptional memory. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=123 SRC="FIGDIR/small/462366v2_ufig1.gif" ALT="Figure 1"> View larger version (30K): org.highwire.dtl.DTLVardef@1d62842org.highwire.dtl.DTLVardef@ed1041org.highwire.dtl.DTLVardef@1c62b43org.highwire.dtl.DTLVardef@1426bcd_HPS_FORMAT_FIGEXP M_FIG C_FIG

molecular biology↗

bioRxiv2020

Loss of mu and delta opioid receptors on neurons expressing dopamine receptor D1 has no effect on reward sensitivity

Opioid signaling controls the activity of the brains reward system. It is involved in signaling the hedonic effects of rewards and also has essential roles in reinforcement and motivational processes. Here, we focused on opioid signaling through mu and delta receptors on dopaminoceptive neurons and evaluated the role these receptors play in reward-driven behaviors. We generated a genetically modified mouse with selective double knockdown of mu and delta opioid receptors in neurons expressing dopamine receptor D1. Selective expression of the transgene was confirmed using immunostaining. Knockdown was validated by measuring the effects of selective opioid receptor agonists on neuronal membrane currents using whole-cell patch clamp recordings. We found that in the nucleus accumbens of control mice, the majority of dopamine receptor D1-expressing neurons were sensitive to a mu or delta opioid agonist. In mutant mice, the response to the delta receptor agonist was blocked, while the effects of the mu agonist were strongly attenuated. Behaviorally, the mice had no obvious impairments. The mutation did not affect sensitivity to the rewarding effects of morphine injections or social contact and had no effect on preference for sweet taste. Knockdown had a moderate effect on motor activity in some of the tests performed, but this effect did not reach statistical significance. Thus, we found that knocking down mu and delta receptors on dopamine receptor D1-expressing cells does not appreciably affect reward-driven behaviors. Highlights- It is well accepted that opioid signaling controls the brains reward system - We generated mutant mice with mu and delta receptor knockdown in D1 neurons - Knockdown made dopaminoceptive neurons insensitive to mu and delta opioid receptor agonists - The mutation did not cause obvious behavioral impairments - The loss of mu and delta receptors on D1 neurons does not affect reward sensitivity

neuroscience↗