Thomas Specht

Expression profiling of microdissected matched prostate cancer samples reveals CD166/MEMD and CD24 as new prognostic markers for patient survival

In order to screen for differentially expressed genes that might be useful in diagnosis or therapy of prostate cancer we have used a custom made Affymetrix GeneChip containing 3950 cDNA fragments. Expression profiles were obtained from 42 matched pairs of mRNAs isolated from microdissected malignant and benign prostate tissues. Applying three different bioinformatic approaches to define differential gene expression, we found 277 differentially expressed genes, of which 98 were identified by all three methods. Fourteen per cent of these genes were not found in other expression studies, which were based on bulk tissue. Resultant candidate genes were further validated by quantitative RT‐PCR, mRNA in situ hybridization and immunohistochemistry. AGR2 was over‐expressed in 89% of prostate carcinomas, but did not have prognostic significance. Immunohistologically detected over‐expression of MEMD and CD24 was identified in 86% and 38.5% of prostate carcinomas respectively, and both were predictive of PSA relapse. Combined marker analysis using MEMD and CD24 expression proved to be an independent prognostic factor (RR = 4.7, p = 0.006) in a Cox regression model, and was also superior to conventional markers. This combination of molecular markers thus appears to allow improved prediction of patient prognosis, but should be validated in larger studies. Copyright

Characterization of recombinant human nicotinamide mononucleotide adenylyl transferase (NMNAT), a nuclear enzyme essential for NAD synthesis

Nicotinamide mononucleotide adenylyl transferase (NMNAT) is an essential enzyme in all organisms, because it catalyzes a key step of NAD synthesis. However, little is known about the structure and regulation of this enzyme. In this study we established the primary structure of human NMNAT. The human sequence represents the first report of the primary structure of this enzyme for an organism higher than yeast. The enzyme was purified from human placenta and internal peptide sequences determined. Analysis of human DNA sequence data then permitted the cloning of a cDNA encoding this enzyme. Recombinant NMNAT exhibited catalytic properties similar to the originally purified enzyme. Human NMNAT (molecular weight 31 932) consists of 279 amino acids and exhibits substantial structural differences to the enzymes from lower organisms. A putative nuclear localization signal was confirmed by immunofluorescence studies. NMNAT strongly inhibited recombinant human poly(ADP‐ribose) polymerase 1, however, NMNAT was not modified by poly(ADP‐ribose). NMNAT appears to be a substrate of nuclear kinases and contains at least three potential phosphorylation sites. Endogenous and recombinant NMNAT were phosphorylated in nuclear extracts in the presence of [γ‐32P]ATP. We propose that NMNATs activity or interaction with nuclear proteins are likely to be modulated by phosphorylation.

5S rRNA Data Bank

In this paper we present the updated version of the compilation of 5S rRNA and 5S rDNA nucleotide sequences. It contains 1622 primary structures of 5S rRNAs and 5S rRNA genes from 888 species. These include 58 archaeal, 427 eubacterial, 34 plastid, nine mitochondrial and 1094 eukaryotic DNA or RNA nucleotide sequences. The sequence entries are divided according to the taxonomic position of the organisms. All individual sequences deposited in the 5S rRNA Database can be retrieved using the WWW-based, taxonomic browser at http://rose.man.poznan.pl/5SData/5SRNA.html++ + or http://www.chemie. fu-berlin.de/fb_chemie/agerdmann/5S_rRNA.html . The files with complete sets of data as well as sequence alignments are available via anonymous ftp.

Does Thermus Represent Another Deep Eubacterial Branching

Summary The complete sequence of a 16S rRNA gene from the extremely thermophilic eubacterium Thermus thermophilus HB8 was determined by sequencing both complementary strands. The sequence has been aligned according to the 16S rRNA compilation by Dams et al. (1988). A phylogenetic tree, based on the alignment of 30 eubacterial and 11 archaebacterial 16S rRNA sequences and with the omission of highly variable regions, was constructed employing a refinement of the neighborliness method. As a result, Thermus thermophilus branches off the line leading to the green non-sulfur bacteria, after Thermotoga the second deepest eubacterial branching so far. The significance of this placement is dicussed.

Compilation of plant 5S ribosomal RNA sequences on RNA and DNA levels

Abstract In this paper we present a compilation of plant 5S ribosomal RNA sequences on both RNA and DNA levels. The comparison of known plant 5S rRNAs and rDNAs shows that most of the sequences of 5S rRNA genes can not be folded into perfect 5S rRNA secondary structure. We propose that an editing mechanism may be involved in correcting of the primary transcripts — repairing of the mismatches — to form mature 5S rRNA molecules.

Compilation of ribosomal 5S ribonucleic acid nucleotide sequences: eukaryotic 5S rRNAs

5S Ribosomal RNA is the smallest RNA component of the ribosomes. Due to relatively simple isolation and sequencing procedures as well as a potential use of the sequence data in evolutionary analyses, the amount of known nucleotide sequences on both RNA and DNA levels was rapidly growing. In this paper we present the updated (March 1996) compilation of eukaryotic 5D rRNA and 5S rDNA sequences.

Evolution of Organisms and Organelles as Studied by Comparative Computer and Biochemical Analyses of Ribosomal 5S RNA Structure

The results documented in this publication demonstrate that for evolutionary studies the ribosomal 5S rRNA is a suitable object for such an investigation and that as many methods as possible should be consulted. In this study the results of biochemical and chemical experiments were combined with those of computer sequence analyses, and they revealed that these methods complement each other nicely. We are currently at a state at which we are able to well define the secondary structures of the 5S rRNAs for eubacteria, organelles, archaebacteria, and eukaryotes and we are even able to propose a secondary structure for a Ur-5S rRNA. It is also clear that in the future the present studies should be continued and extended in such a way that the tertiary structures of these molecules will become known.