A newly identifiedMinute locus,M(2)32D, encodes the ribosomal protein L9 inDrosophila melanogaster

A gene encoding a ubiquitously expressed mRNA inDrosophila melanogaster was isolated and identified as the gene for ribosomal protein L9 (rpL9) by its extensive sequence homology to the corresponding gene from rat. TherpL9 gene is localized in polytene region 32D where two independent P element insertions flanking the locus are available. Remobilization of either P element generated lines with a typicalMinute phenotype, e.g. thin and short bristles, prolonged development, and female semisterility in heterozygotes as well as homozygous lethality. All these characteristics can be rescued when a 3.9 kb restriction fragment containing therpL9 gene is reintroduced by P element-mediated germline transformation. This result confirms thatM(2)32D codes for ribosomal protein L9.     

cDNA, genomic sequence cloning and overexpression of ribosomal protein gene L9 (rpL9) of the giant panda (Ailuropoda melanoleuca)

The ribosomal protein L9 (RPL9), a component of the large subunit of the ribosome, has an unusual structure, comprising two compact globular domains connected by an α-helix; it interacts with 23 S rRNA. To obtain information about rpL9 of Ailuropoda melanoleuca (the giant panda) we designed primers based on the known mammalian nucleotide sequence. RT-PCR and PCR strategies were employed to isolate cDNA and the rpL9 gene from A. melanoleuca; these were sequenced and analyzed. We overexpressed cDNA of the rpL9 gene in Escherichia coli BL21. The cloned cDNA fragment was 627 bp in length, containing an open reading frame of 579 bp. The deduced protein is composed of 192 amino acids, with an estimated molecular mass of 21.86 kDa and an isoelectric point of 10.36. The length of the genomic sequence is 3807 bp, including six exons and five introns. Based on alignment analysis, rpL9 has high similarity among species; we found 85% agreement of DNA and amino acid sequences with the other species that have been analyzed. Based on topology predictions, there are two N-glycosylation sites, five protein kinase C phosphorylation sites, one casein kinase II phosphorylation site, two tyrosine kinase phosphorylation sites, three N-myristoylation sites, one amidation site, and one ribosomal protein L6 signature 2 in the L9 protein of A. melanoleuca. The rpL9 gene can be readily expressed in E. coli; it fuses with the N-terminal GST-tagged protein, giving rise to the accumulation of an expected 26.51-kDa polypeptide, which is in good agreement with the predicted molecular weight. This expression product could be used for purification and further study of its function.     

The gene family encoding the mouse ribosomal protein L32 contains a uniquely expressed intron-containing gene and an unmutated processed gene

The family of approximately 16 genes encoding the mouse ribosomal protein L32 has been characterized by an analysis of a representative set of genomic clones. Surprisingly, this family contains only a single expressed intron-containing gene. This gene, termed rpL32 , has been completely sequenced and found to possess certain novel features including the presence in two of its introns of a sequence with high homology to the 5' end of U1 snRNA and a 5' terminal region exceptionally rich in pyrimidines. Most of the other members of the L32 family appear to be processed genes, some of which are identical or very similar to the rpL32 gene, except for the lack of introns. One unmutated gene was found to be integrated 28 nucleotides downstream of a canonical TATA box. However, despite this feature, the gene does not seem to be expressed, as judged by its extent of methylation compared to the expressed rpL32 gene.     

Suppression of the ribosomal L2 gene reveals a novel mechanism for stress adaptation in soybean

Pseudomonas syringae pv. glycinea bacteria or zoospores of the fungus Phytophthora sojae were used to trigger a hypersensitive reaction (HR) in cell cultures of soybean (Glycine max [L.] Merr. cv. Williams 82). During a screen for genes that show an altered expression as a response to dying neighbour cells we have identified a gene fragment that is specifically but transiently down-regulated in an HR. The corresponding cDNA codes for the ribosomal protein L2 (rpL2) of 80S ribosomes, which is essential for the peptidyl-transferase activity. Two gene copies of rpL2 exist in soybean and both genes are transcribed. The temporary down-regulation of the rpL2 genes is followed by a transient block in the synthesis of new proteins as visualised by pulse-labelling experiments using ³⁵S-amino acids. The same basic phenomenon was also found after treatment of soybean cells with other stress-causing compounds such as elicitors or heavy metals. It is suggested that the transient block in protein synthesis allows a more rapid depletion of, for example, signal molecules with a short half-life time and thus leads to a faster adaptation of the cellular protein inventory to the new environmental conditions. Received: 10 May 2000 / Accepted: 21 July 2000     

Cloning and sequencing of ribosomal protein L27a and a gene similar to human GS1 in Drosophila

Two closely linked genes were identified and characterized in the 24F region on the left arm of chromosome 2 in Drosophila. One cDNA predicts a protein of 231 amino acids, with a molecular mass of 25.7 kDa. The predicted amino-acid sequence of this protein is 47.2% identical to that of the previously reported human GS1 protein, which is encoded by a gene that is of interest because it is one of the few X-linked genes that escapes X-inactivation. We have accordingly named our gene GS1like (GS1l). The second cDNA begins 383 bp proximal to the first. This cDNA encodes a protein of a predicted 149 amino acids and a molecular mass of 17.0 kDa. This protein represents a homolog of ribosomal protein L27a; thus, we have named the gene RpL27a. This gene might be responsible for the Minute mutation located at 24F. An rpL27a gene was previously localized to 87F/88A; thus, this gene might be present in two locations in Drosophila.     

Phosphorylation of ribosomal proteins induced by auxins in maize embryonic tissues

The effect of auxin on ribosomal protein phosphorylation of germinating maize (Zea mays) tissues was investigated. Two-dimensional gel electrophoresis and autoradiography of [³²P] ribosomal protein patterns for natural and synthetic auxin-treated tissues were performed. Both the rate of ³²P incorporation and the electrophoretic patterns were dependent on ³²P pulse length, suggesting that active protein phosphorylation-dephosphorylation occurred in small and large subunit proteins, in control as well as in auxin-treated tissues. The effect of ribosomal protein phosphorylation on in vitro translation was tested. Measurements of poly(U) translation rates as a function of ribosome concentration provided apparent K_m values significantly different for auxin-treated and nontreated tissues. These findings suggest that auxin might exert some kind of translational control by regulating the phosphorylated status of ribosomal proteins.     

Studies on the Assembly Characteristics of Large Subunit Ribosomal Proteins in S. cerevisae

During the assembly process of ribosomal subunits, their structural components, the ribosomal RNAs (rRNAs) and the ribosomal proteins (r-proteins) have to join together in a highly dynamic and defined manner to enable the efficient formation of functional ribosomes. In this work, the assembly of large ribosomal subunit (LSU) r-proteins from the eukaryote S. cerevisiae was systematically investigated. Groups of LSU r-proteins with specific assembly characteristics were detected by comparing the protein composition of affinity purified early, middle, late or mature LSU (precursor) particles by semi-quantitative mass spectrometry. The impact of yeast LSU r-proteins rpL25, rpL2, rpL43, and rpL21 on the composition of intermediate to late nuclear LSU precursors was analyzed in more detail. Effects of these proteins on the assembly states of other r-proteins and on the transient LSU precursor association of several ribosome biogenesis factors, including Nog2, Rsa4 and Nop53, are discussed.