Tet controls axon guidance in early brain development through glutamatergic signaling.

Mutations in ten-eleven translocation (TET) proteins are associated with human neurodevelopmental disorders. We find a function of Tet in regulating early brain development. The Tet DNA-binding domain () is required for axon guidance in the mushroom body (MB).

Tet-dependent 5-hydroxymethyl-Cytosine modification of mRNA regulates axon guidance genes in Drosophila.

Modifications of mRNA, especially methylation of adenosine, have recently drawn much attention. The much rarer modification, 5-hydroxymethylation of cytosine (5hmC), is not well understood and is the subject of this study. Vertebrate Tet proteins are 5-methylcytosine (5mC) hydroxylases and catalyze the transition of 5mC to 5hmC in DNA.

Tet Controls Axon Guidance in Early Brain Development through Glutamatergic Signaling.

Mutations in human TET proteins have been found in individuals with neurodevelopmental disorders. Here we report a new function of Tet in regulating early brain development. We found that mutation in the Tet DNA-binding domain ( ) resulted in axon guidance defects in the mushroom body (MB).

Tet-dependent 5-hydroxymethyl-Cytosine modification of mRNA regulates the axon guidance genes and in .

Modifications of mRNA, especially methylation of adenosine, have recently drawn much attention. The much rarer modification, 5-hydroxymethylation of cytosine (5hmC), is not well understood and is the subject of this study. Vertebrate Tet proteins are 5-methylcytosine (5mC) hydroxylases enzymes catalyzing the transition of 5mC to 5hmC in DNA and have recently been shown to have the same function in messenger RNAs in both vertebrates and in Drosophila.

Tet-dependent 5-hydroxymethyl-Cytosine modification of mRNA regulates axon guidance genes in Drosophila.

Modifications of mRNA, especially methylation of adenosine, have recently drawn much attention. The much rarer modification, 5-hydroxymethylation of cytosine (5hmC), is not well understood and is the subject of this study. Vertebrate Tet proteins are 5-methylcytosine (5mC) hydroxylases and catalyze the transition of 5mC to 5hmC in DNA.

Expression and functional analysis of various structural domains of tobacco topoisomerase II: To understand the mechanistic insights of plant type II topoisomerases.

In contrast to bacterial, yeast and animal systems, topoisomerases (topo) from plants have not been well studied. In this report, we generated four truncated topoisomerase II (Topo II) cDNA fragments encoding different functional domains of Nicotiana tabacum topo II (NtTopoII). Each of these recombinant polypeptides was expressed alone or in combination in temperature-sensitive topoisomerase II yeast mutants.

Over-expression of Topoisomerase II Enhances Salt Stress Tolerance in Tobacco.

Topoisomerases are unique enzymes having an ability to remove or add DNA supercoils and untangle the snarled DNA. They can cut, shuffle, and religate DNA strands and remove the torsional stress during DNA replication, transcription or recombination events. In the present study, we over-expressed topoisomerase II (TopoII) in tobacco (Nicotiana tabaccum) and examined its role in growth and development as well as salt (NaCl) stress tolerance.

Promoter-Terminator Gene Loops Affect Alternative 3'-End Processing in Yeast.

Many eukaryotic genes undergo alternative 3'-end poly(A)-site selection producing transcript isoforms with 3'-UTRs of different lengths and post-transcriptional fates. Gene loops are dynamic structures that juxtapose the 3'-ends of genes with their promoters. Several functions have been attributed to looping, including memory of recent transcriptional activity and polarity of transcription initiation.

Detection of short-range chromatin interactions by chromosome conformation capture (3C) in yeast.

We describe a modified 3C ("chromosome conformation capture") protocol for detection of transient, short-range chromatin interactions in the yeast Saccharomyces cerevisiae. 3C was initially described by Job Dekker and involves formaldehyde cross-linking to stabilize transient chromatin interactions, followed by restriction digestion, ligation, and locus-specific PCR. As such, 3C reveals complex three-dimensional interactions between distal genetic elements within intact cells at high resolution.

DNA looping facilitates targeting of a chromatin remodeling enzyme.

ATP-dependent chromatin remodeling enzymes are highly abundant and play pivotal roles regulating DNA-dependent processes. The mechanisms by which they are targeted to specific loci have not been well understood on a genome-wide scale. Here, we present evidence that a major targeting mechanism for the Isw2 chromatin remodeling enzyme to specific genomic loci is through sequence-specific transcription factor (TF)-dependent recruitment.

The interaction of Pcf11 and Clp1 is needed for mRNA 3'-end formation and is modulated by amino acids in the ATP-binding site.

Polyadenylation of eukaryotic mRNAs contributes to stability, transport and translation, and is catalyzed by a large complex of conserved proteins. The Pcf11 subunit of the yeast CF IA factor functions as a scaffold for the processing machinery during the termination and polyadenylation of transcripts. Its partner, Clp1, is needed for mRNA processing, but its precise molecular role has remained enigmatic.

Control of eukaryotic gene expression: gene loops and transcriptional memory.

Gene loops are dynamic structures that juxtapose promoter–terminator regions of Pol II-transcribed genes. Although first described in yeast, gene loops have now been identified in yeast and mammalian cells. Looping requires components of the transcription preinitiation complex, the pre-mRNA 30-end processing machinery, and subunits of the nuclear pore complex.

A physiological role for gene loops in yeast.

DNA loops that juxtapose the promoter and terminator regions of RNA polymerase II-transcribed genes have been identified in yeast and mammalian cells. Loop formation is transcription-dependent and requires components of the pre-mRNA 3'-end processing machinery. Here we report that looping at the yeast GAL10 gene persists following a cycle of transcriptional activation and repression.

Detection of gene loops by 3C in yeast.

"Chromosome conformation capture" (3C) is a powerful method to detect physical interaction between any two genomic loci. 3C involves formaldehyde crosslinking to stabilize transient interactions, followed by restriction digestion, ligation and locus-specific PCR. Accordingly, 3C reveals complex three-dimensional interactions between distal genetic elements within intact cells at high resolution.

The essential N terminus of the Pta1 scaffold protein is required for snoRNA transcription termination and Ssu72 function but is dispensable for pre-mRNA 3'-end processing.

Saccharomyces cerevisiae Pta1 is a component of the cleavage/polyadenylation factor (CPF) 3'-end processing complex and functions in pre-mRNA cleavage, poly(A) addition, and transcription termination. In this study, we investigated the role of the N-terminal region of Pta1 in transcription and processing. We report that a deletion of the first 75 amino acids (pta1-Delta75) causes thermosensitive growth, while the deletion of an additional 25 amino acids is lethal.

A transcription-independent role for TFIIB in gene looping.

Recent studies demonstrated the existence of gene loops that juxtapose the promoter and terminator regions of genes with exceptionally long ORFs in yeast. Here we report that looping is not idiosyncratic to long genes but occurs between the distal ends of genes with ORFs as short as 1 kb. Moreover, looping is dependent upon the general transcription factor TFIIB: the E62K (glutamic acid 62 --> lysine) form of TFIIB adversely affects looping at every gene tested, including BLM10, SAC3, GAL10, SEN1, and HEM3.

Regulation of transcript elongation through cooperative and ordered recruitment of cofactors.

We studied the regulation of murine CD80, a gene whose basal transcriptional status was characterized by the presence of a stalled RNA polymerase II complex on the promoter-proximal region. Stimulus-induced activation of productive elongation involved a complex interplay of regulated events that included a synergy between ordered cofactor recruitment. This cascade of recruitments was initiated through the engagement of transcription factor NF-kappaB, leading to the temporal association of histone acetyltransferases and the consequent selective acetylation of a transcription start site downstream nucleosome.

Transcription regulation from a TATA and INR-less promoter: spatial segregation of promoter function.

The mode of regulation of class II genes that lack the known core promoter elements is presently unclear. Here, we studied one such example, the murine CD80 gene. An unusual mechanism was revealed wherein the pre-initiation complex (PIC) first assembled on an upstream, NF-kappaB enhancer element.