Seema Irani

Bio: Seema Irani is an academic researcher from University of Texas at Austin. The author has contributed to research in topics: RNA polymerase II & Phosphorylation. The author has an hindex of 8, co-authored 19 publications receiving 153 citations.

Identification of the Formycin A Biosynthetic Gene Cluster from Streptomyces kaniharaensis Illustrates the Interplay between Biological Pyrazolopyrimidine Formation and de Novo Purine Biosynthesis.

Abstract: Formycin A is a potent purine nucleoside antibiotic with a C-glycosidic linkage between the ribosyl moiety and the pyrazolopyrimidine base Herein, a cosmid is identified from the Streptomyces kaniharaensis genome library that contains the for gene cluster responsible for the biosynthesis of formycin Subsequent gene deletion experiments and in vitro characterization of the forBCH gene products established their catalytic functions in formycin biosynthesis Results also demonstrated that PurH from de novo purine biosynthesis plays a key role in pyrazolopyrimidine formation during biosynthesis of formycin A The participation of PurH in both pathways represents a good example of how primary and secondary metabolism are interlinked

Enzyme-mediated depletion of serum l-Met abrogates prostate cancer growth via multiple mechanisms without evidence of systemic toxicity.

Abstract: Extensive studies in prostate cancer and other malignancies have revealed that l-methionine (l-Met) and its metabolites play a critical role in tumorigenesis. Preclinical and clinical studies have demonstrated that systemic restriction of serum l-Met, either via partial dietary restriction or with bacterial l-Met-degrading enzymes exerts potent antitumor effects. However, administration of bacterial l-Met-degrading enzymes has not proven practical for human therapy because of problems with immunogenicity. As the human genome does not encode l-Met-degrading enzymes, we engineered the human cystathionine-γ-lyase (hMGL-4.0) to catalyze the selective degradation of l-Met. At therapeutically relevant dosing, hMGL-4.0 reduces serum l-Met levels to >75% for >72 h and significantly inhibits the growth of multiple prostate cancer allografts/xenografts without weight loss or toxicity. We demonstrate that in vitro, hMGL-4.0 causes tumor cell death, associated with increased reactive oxygen species, S-adenosyl-methionine depletion, global hypomethylation, induction of autophagy, and robust poly(ADP-ribose) polymerase (PARP) cleavage indicative of DNA damage and apoptosis.

Mapping the Phosphorylation Pattern of Drosophila melanogaster RNA Polymerase II Carboxyl-Terminal Domain Using Ultraviolet Photodissociation Mass Spectrometry.

Abstract: Phosphorylation of the C-terminal domain of RNA polymerase II (CTD) plays an essential role in eukaryotic transcription by recruiting transcriptional regulatory factors to the active polymerase. However, the scarcity of basic residues and repetitive nature of the CTD sequence impose a huge challenge for site-specific characterization of phosphorylation, hindering our understanding of this crucial biological process. Herein, we apply LC-UVPD-MS methods to analyze post-translational modification along native sequence CTDs. Application of our method to the Drosophila melanogaster CTD reveals the phosphorylation pattern of this model organism for the first time. The divergent nature of fly CTD allows us to derive rules defining how flanking residues affect phosphorylation choice by CTD kinases. Our data support the use of LC-UVPD-MS to decipher the CTD code and determine rules that program its function.

Tyr1 phosphorylation promotes phosphorylation of Ser2 on the C-terminal domain of eukaryotic RNA polymerase II by P-TEFb

Abstract: DNA contains the instructions for making proteins, which build and maintain our cells. So that the information encoded in DNA can be used, a molecular machine called RNA polymerase II makes copies of specific genes. These copies, in the form of a molecule called RNA, convey the instructions for making proteins to the rest of the cell. To ensure that RNA polymerase II copies the correct genes at the correct time, a group of regulatory proteins are needed to control its activity. Many of these proteins interact with RNA polymerase II at a region known as the C-terminal domain, or CTD for short. For example, before RNA polymerase can make a full copy of a gene, a small molecule called a phosphate group must first be added to CTD at specific units known as Ser2. The regulatory protein P-TEFb was thought to be responsible for phosphorylating Ser2. However, it was previously not known how P-TEFb added this phosphate group, and why it did not also add phosphate groups to other positions in the CTD domain that are structurally similar to Ser2. To investigate this, Mayfield, Irani et al. mixed the CTD domain with different regulatory proteins, and used various biochemical approaches to examine which specific positions of the domain had phosphate groups attached. These experiments revealed a previously unknown aspect of P-TEFb activity: its specificity for Ser2 increased dramatically if a different regulatory protein first added a phosphate group to a nearby location in CTD. This additional phosphate group directed P-TEFb to then add its phosphate specifically at Ser2. To confirm the activity of this mechanism in living human cells, Mayfield, Irani et al. used a drug that prevented the first phosphate from being added. In the drug treated cells, RNA polymerase II was found more frequently ‘stalled’ at positions on the DNA just before a gene starts. This suggests that living cells needs this two-phosphate code system in order for RNA polymerase II to progress and make copies of specific genes. These results are a step forward in understanding the complex control mechanisms cells use to make proteins from their DNA. Moreover, the model presented here – one phosphate addition priming a second specific phosphate addition – provides a template that may underlie similar regulatory processes.

Integrative Genomics Viewer

Abstract: Rapid improvements in sequencing and array-based platforms are resulting in a flood of diverse genome-wide data, including data from exome and whole-genome sequencing, epigenetic surveys, expression profiling of coding and noncoding RNAs, single nucleotide polymorphism (SNP) and copy number profiling, and functional assays. Analysis of these large, diverse data sets holds the promise of a more comprehensive understanding of the genome and its relation to human disease. Experienced and knowledgeable human review is an essential component of this process, complementing computational approaches. This calls for efficient and intuitive visualization tools able to scale to very large data sets and to flexibly integrate multiple data types, including clinical data. However, the sheer volume and scope of data pose a significant challenge to the development of such tools.

Decision-making in structure solution using Bayesian estimates of map quality: the PHENIX autosol wizard

Abstract: Ten measures of experimental electron-density-map quality are examined and the skewness of electron density is found to be the best indicator of actual map quality. A Bayesian approach to estimating map quality is developed and used in the PHENIX AutoSol wizard to make decisions during automated structure solution.

Top-Down Proteomics: Ready for Prime Time?

Abstract: In the post-genomics era, the study of proteins is critical for understanding cellular functions at the molecular level.1-5 Beyond the genetic code, the human proteome is greatly diversified by various genetic variations, alternative splicing of RNA transcripts, and post-translational modifi-cations (PTMs).6,7 In 2013, the term “proteoform” was designated to describe “all of the different molecular forms in which the protein product of a single gene can be found”,6 clearing up the confusion in nomenclature and joining research efforts to develop methodologies for pro-teoform characterization. Top-down proteomics, which analyzes intact proteins without digestion, has proved to be a premier technology for global and comprehensive analy-sis of proteoforms.4,8,9 The top-down approach retains in-tact protein mass information, providing a “bird’s-eye” view of the proteome and allowing for identification of novel proteoforms, in-depth sequence characterization, and quantification of disease-associated PTMs.4,8,9

Ultraviolet Photodissociation Mass Spectrometry for Analysis of Biological Molecules.

Abstract: The development of new ion-activation/dissociation methods continues to be one of the most active areas of mass spectrometry owing to the broad applications of tandem mass spectrometry in the identification and structural characterization of molecules. This Review will showcase the impact of ultraviolet photodissociation (UVPD) as a frontier strategy for generating informative fragmentation patterns of ions, especially for biological molecules whose complicated structures, subtle modifications, and large sizes often impede molecular characterization. UVPD energizes ions via absorption of high-energy photons, which allows access to new dissociation pathways relative to more conventional ion-activation methods. Applications of UVPD for the analysis of peptides, proteins, lipids, and other classes of biologically relevant molecules are emphasized in this Review.