Tip off the HAT– Epigenetic control of learning and memory by Drosophila Tip60

Disruption of epigenetic gene control mechanisms involving histone acetylation in the brain causes cognitive impairment, a debilitating hallmark of most neurodegenerative disorders. Histone acetylation regulates cognitive gene expression via chromatin packaging control in neurons. Unfortunately, the histone acetyltransferases (HATs) that generate such neural epigenetic signatures and their mechanisms of action remain unclear. Our recent findings provide insight into this question by demonstrating that Tip60 HAT action is critical for morphology and function of the mushroom body (MB), the learning and memory center in the Drosophila brain. We show that Tip60 is robustly produced in MB Kenyon cells and extending axonal lobes and that targeted MB Tip60 HAT loss results in axonal outgrowth disruption. Functional consequences of loss and gain of Tip60 HAT levels in the MB are evidenced by defects in memory. Tip60 ChIP-Seq analysis reveals enrichment for genes that function in cognitive processes and accordingly, key genes representing these pathways are misregulated in the Tip60 HAT mutant fly brain. Remarkably, increasing levels of Tip60 in the MB rescues learning and memory deficits resulting from Alzheimer''s disease associated amyloid precursor protein (APP) induced neurodegeneration. Our studies highlight the potential of HAT activators as a therapeutic option for cognitive disorders.     

Elucidation of molecular mechanism of stability of the heme-regulated eIF2α kinase upon binding of its ligand,hemin in its catalytic kinase domain

The eIF2α kinase activity of the heme-regulated inhibitor (HRI) is regulated by heme which makes it a unique member of the family of eIF2α kinases. Since heme concentrations create an equilibrium for the kinase to be active/inactive, it becomes important to study the heme binding effects upon the kinase and understanding its mechanism of functionality. In the present study, we report the thermostability achieved by the catalytic kinase domain of HRI (HRI.CKD) upon ligand (heme) binding. Our CD data demonstrates that the HRI.CKD retains its secondary structure at higher temperatures when it is in ligand bound state. HRI.CKD when incubated with hemin loses its monomeric state and attains a higher order oligomeric form resulting in its stability. The HRI.CKD fails to refold into its native conformation upon mutation of H377A/H381A, thereby confirming the necessity of these His residues for correct folding, stability, and activity of the kinase. Though our in silico study demonstrated these His being the ligand binding sites in the kinase insert region, the spectra-based study did not show significant difference in heme affinity for the wild type and His mutant HRI.CKD.     

Thlaspi s.str. (Brassicaceae) versus Thlaspi s.l.: morphological and anatomical characters in the light of ITS nrDNA sequence data

 Nuclear ribosomal (ITS) DNA sequence data from representatives of eleven out of 12 genera previously included in Thlaspi sensu lato were analyzed to elucidate relationships within the former genus Thlaspi sensu lato. Sequences from Thlaspi segregates Noccaea, Masmenia, Callothlaspi, Kotschyella, Microthlaspi, and Noccidium were added to the existing data sets, and only some Thlaspi sensu lato segregates formed a monophyletic group. The recently renamed genus Pseudosempervivum, formerly part of the genus Cochlearia, has been shown to be closely related to this group as well. Microthlaspi consists of three independent lineages of which Microthlaspi umbellatum is closely related to Neurotropis. The Thlaspi s.str. taxa including the type species T. arvense, are closely related to Peltaria and Alliaria and they represent a monophyletic group. Kotschyella and Noccidium, which were also integrated into Thlaspi s.l. are not closely related to any Thlaspi lineage. Therefore, we suggest major taxonomical changes which are not in agreement with concepts based on morphological data. Our ITS based phylogeny demonstrates that subtribe Thlaspidinae comprises some additional genera such as Teesdalia, Peltaria, and Alliaria, the latter two even previously classified in other tribes than tribe Lepidieae. Received August 15, 2000 Accepted January 30, 2001     

Basal euteleostean relationships: a mitogenomic perspective on the phylogenetic reality of the "Protacanthopterygii"

Higher-level relationships of the basal Euteleostei (=Protacanthopterygii) are so complex and controversial that at least nine different morphology-based phylogenetic hypotheses have been proposed during the last 30 years. Relationships of the Protacanthopterygii were investigated using mitochondrial genomic (mitogenomic) data from 34 purposefully chosen species (data for 12 species being newly determined during the study) that fully represented major basal euteleostean lineages and some basal teleosts plus neoteleosts as outgroups. Unweighted and weighted maximum parsimony (MP) and maximum likelihood (ML) analyses were conducted with the data set that comprised concatenated nucleotide sequences from 12 protein-coding genes (excluding the ND6 gene and 3rd codon positions) and 22 transfer RNA (tRNA) genes (stem regions only) from the 34 species. The resultant trees were well resolved and largely congruent, with most internal branches being supported by high statistical values. Monophyly of the protacanthopterygians was confidently rejected by the mitogenomic data. Of the five major monophyletic groups that received high statistical support within the protacanthopterygians, a clade comprising members of the alepocephaloids was unexpectedly nested within the Otocephala, sister-group of the euteleosts. The remaining four major monophyletic groups, on the other hand, occupied phylogenetic positions intermediate between the otocephalans and neoteleosts, with a clade comprising esociforms + salmoniforms being more basal to the argentinoids and osmeroids. Although interrelationships of the latter two clades (argentinoids and osmeroids) with the neoteleosts remained ambiguous, the present results indicated explicitly that the protacanthopterygians as currently defined merely represent a collective, polyphyletic group of the basal euteleosts, located between the basal teleosts (elopomorphs and below) and neoteleosts (stomiiforms and above).     

Drosophila heparan sulfate, a novel design

Heparan sulfate (HS) proteoglycans play critical roles in a wide variety of biological processes such as growth factor signaling, cell adhesion, wound healing, and tumor metastasis. Functionally important interactions between HS and a variety of proteins depend on specific structural features within the HS chains. The fruit fly (Drosophila melanogaster) is frequently applied as a model organism to study HS function in development. Previous structural studies of Drosophila HS have been restricted to disaccharide composition, without regard to the arrangement of saccharide domains typically found in vertebrate HS. Here, we biochemically characterized Drosophila HS by selective depolymerization with nitrous acid. Analysis of the generated saccharide products revealed a novel HS design, involving a peripheral, extended, presumably single, N-sulfated domain linked to an N-acetylated sequence contiguous with the linkage to core protein. The N-sulfated domain may be envisaged as a heparin structure of unusually low O-sulfate content.