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A S Brodsky

Publications and source records attributed to A S Brodsky.

4 recordsLinked to original sources
PubMed2000

Pre-mRNA processing factors are required for nuclear export.

RNA export from the nucleus is thought to be linked to proper processing and packaging into ribonucleoprotein protein complexes. A system to observe mRNA nuclear export in living yeast cells was developed by fusing the U1A RNA-binding protein to the green fluorescent protein to follow specific mRNAs with U1A hairpins engineered into them. RNAs encoding Rpl25, Pgk1, and Ssa4 were examined for the effects of 3' UTRs, introns, RNA processing factors, nucleoporins, and transport factors on their export. All accumulated in the nucleus in mutants affecting components of the nuclear export machinery and certain nucleoporins. However, under conditions of stress, PGK1 and RPL25 transcripts accumulate in the nucleus whereas SSA4 RNA is exported. Moreover, when export is blocked, only RNAs containing the ASH1 3' UTR accumulated in the nucleolus. Mutations in the splicing machinery selectively blocked export of only intron-containing RNAs. Mutations in RNA14, RNA15, and PAP1, which encode factors important for 3' processing, also blocked export of all RNAs, including SSA4, thereby linking export to the process of polyadenlyation. Taken together, these data graphically display the connections between mRNA processing and nuclear export.

3' Untranslated Regions↗
PubMed1998

NMR evidence for a base triple in the HIV-2 TAR C-G.C+ mutant-argininamide complex.

Formation of a specific complex between the HIV Tat protein and the small RNA element TAR is critical for activation of viral transcription. A model complex for this interaction composed of HIV-2 TAR and the amide derivative of arginine has been developed to study how Tat and TAR interact specifically. We have previously determined a high resolution NMR structure of the HIV-2 TAR-argininamide complex. The argininamide guanidium group hydrogen bonds to the major groove face of G26 and is stacked between U23 and A22, forming an arginine sandwich. This structure also provided evidence for formation of a U38-A27.U38 base triple, as U23 is positioned in the major groove within hydrogen bonding distance to A27. However, the expected U23 imino proton was not observed, preventing unambiguous identification of the base triple. Previous work on an isomorphic C38-G27.C23+ base triple mutant of the three base bulge HIV-1 TAR-argininamide complex demonstrated that the base triple is required for specific argininamide binding. Here we investigate the same C38-G27.C23+ base triple mutant in the context of two base bulge HIV-2 TAR. The improved NMR spectral properties of HIV-2 TAR allowed observation of the C23 amino and imino protons for the first time, providing direct evidence that a hydrogen bonding interaction is occurring. The NOEs observed correspond to those observed in the high resolution structure of the HIV-2 TAR-argininamide complex, confirming that a base triple is an important feature of the TAR-argininamide interaction.

Arginine↗
PubMed1997

Solution structure of the HIV-2 TAR-argininamide complex.

The trans-activating region (TAR) RNA-Tat protein interaction is important for activation of transciption in the human immunodeficiency virus (HIV). A model complex for this interaction composed of the two base bulge HIV-2 TAR and the amide derivative of arginine was studied by multidimensional heteronuclear NMR. Because of the improved spectral properties of the HIV-2 TAR complex, a larger number of NOEs in the bulge region were observed than in earlier studies of the HIV-1 TAR-argininamide complex. A total of 681 NOE distance restraints were collected and used to determine the solution structure of the HIV-2 TAR-argininamide complex. As observed in the previously proposed model from this lab, the two A-form stems co-axially stack and the critical U23 and the argininamide are located in the major groove. Model calculations including non-experimental restraints indicate that U23 is within hydrogen bonding distance to A27 consistent with the formation of a U x A x U base-triple. Base-triple formation helps open the major groove to increase the accessibility of G26 to hydrogen bond donors from the guanidinium group of argininamide. Argininamide binding is stabilized by stacking of the guanidinium group between the bases of A22 and U23, forming an argininamide sandwich.

Arginine↗