scholarly journals Four Ia invariant chain forms derive from a single gene by alternate splicing and alternate initiation of transcription/translation.

1987 ◽  
Vol 166 (2) ◽  
pp. 444-460 ◽  
Author(s):  
D M O'Sullivan ◽  
D Noonan ◽  
V Quaranta
Keyword(s):  
Single Gene ◽  
Structural Basis ◽  
Invariant Chain ◽  
Nucleotide Sequencing ◽  
S1 Nuclease ◽  
Initiation Of Translation ◽  
Associated Proteins ◽  
Alternatively Spliced ◽  
Translational Start ◽  
Genomic Regions

We determined the structural basis for the presence of electrophoretically-distinct, antigenically-related forms of invariant chains in Ia oligomers, and established the mechanisms by which they can be expressed from a single gene. S1 nuclease protection assays indicated that, in B cells, transcription of this gene initiates at a minimum of three sites. Thus, unlike previously thought, invariant chain mRNAs have heterogeneous 5' untranslated segments that may differentially affect initiation of translation. Further, restriction mapping and nucleotide sequencing of cDNAs revealed two kinds of invariant chain mRNAs differing by an internal coding segment of 192 bp. This segment represents an alternatively spliced exon, as demonstrated by nucleotide sequencing of corresponding genomic regions. The exon (exon X) encodes a cysteine-rich stretch of 64 amino acids near the COOH terminus that displays a striking and surprising homology to an internal amino acid repeat of thyroglobulin, suggesting an evolutionary mechanism of exon shuffling. Transient expression of cDNAs indicated that both types of alternatively spliced mRNAs contain two in-frame AUGs functioning as alternate start sites for translation. Thus, transfections with exon X-lacking cDNAs resulted in the expression of Mr 33,000 and 31,000 proteins, detected by immunoprecipitation with anti-invariant chain antisera, and identical by two-dimensional gel (2-D) analyses to the B cell invariant-chain forms gamma 1 (Mr 31,000), gamma 2, and gamma 3 (Mr 33,000). Similarly, exon X-containing cDNAs expressed Mr 43,000 and 41,000 proteins, also identical by 2-D migration to Ia-associated proteins. Thus, human Ia molecules contain four forms of invariant chain of closely related but nonidentical primary structure that are generated from a single gene by a complex pattern of alternate transcriptional start, exon splicing, and translational start.

scholarly journals Fine-Mapping of Sorghum Stay-Green QTL on Chromosome10 Revealed Genes Associated with Delayed Senescence

Genes ◽  
2020 ◽  
Vol 11 (9) ◽  
pp. 1026 ◽  
Author(s):  
K. N. S. Usha Kiranmayee ◽  
C. Tom Hash ◽  
S. Sivasubramani ◽  
P. Ramu ◽  
Bhanu Prakash Amindala ◽  
...  
Keyword(s):  
Drought Tolerance ◽  
Fine Mapping ◽  
Mapping Population ◽  
Single Gene ◽  
Genotyping By Sequencing ◽  
Introgression Line ◽  
Genomic Region ◽  
Stay Green ◽  
Delayed Senescence ◽  
Genomic Regions

This study was conducted to dissect the genetic basis and to explore the candidate genes underlying one of the important genomic regions on an SBI-10 long arm (L), governing the complex stay-green trait contributing to post-flowering drought-tolerance in sorghum. A fine-mapping population was developed from an introgression line cross—RSG04008-6 (stay-green) × J2614-11 (moderately senescent). The fine-mapping population with 1894 F2 was genotyped with eight SSRs and a set of 152 recombinants was identified, advanced to the F4 generation, field evaluated with three replications over 2 seasons, and genotyped with the GBS approach. A high-resolution linkage map was developed for SBI-10L using 260 genotyping by sequencing—Single Nucleotide Polymorphism (GBS–SNPs). Using the best linear unpredicted means (BLUPs) of the percent green leaf area (%GL) traits and the GBS-based SNPs, we identified seven quantitative trait loci (QTL) clusters and single gene, mostly involved in drought-tolerance, for each QTL cluster, viz., AP2/ERF transcription factor family (Sobic.010G202700), NBS-LRR protein (Sobic.010G205600), ankyrin-repeat protein (Sobic.010G205800), senescence-associated protein (Sobic.010G270300), WD40 (Sobic.010G205900), CPK1 adapter protein (Sobic.010G264400), LEA2 protein (Sobic.010G259200) and an expressed protein (Sobic.010G201100). The target genomic region was thus delimited from 15 Mb to 8 genes co-localized with QTL clusters, and validated using quantitative real-time (qRT)–PCR.

scholarly journals The pyrimidine/purine-biased region of the epidermal growth factor receptor gene is sensitive to S1 nuclease and may form an intramolecular triplex

1990 ◽  
Vol 268 (1) ◽  
pp. 175-180 ◽  
Author(s):  
M Kato ◽  
J Kudoh ◽  
N Shimizu
Keyword(s):  
Epidermal Growth Factor Receptor ◽  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Transcription Initiation ◽  
Receptor Gene ◽  
Growth Factor Receptor ◽  
Structural Basis ◽  
S1 Nuclease ◽  
Single Stranded Region ◽  
Epidermal Growth

The pyrimidine/purine-biased region located upstream of the EGF (epidermal growth factor) receptor gene transcription initiation sites was sensitive to S1 nuclease when under superhelical tension. The structural basis of this specific reactivity to S1 nuclease was probed by the use of diethyl pyrocarbonate. The patterns of modification suggested that the H-form proposed by Mirkin, Lyamichev, Drushlyak, Dobrynin, Filippov & Frank-Kamenetskii [Nature (London) (1987) 330, 495-497], which includes an intramolecular triplex and a single-stranded region, was the most plausible model for the sequence tested. The results of dimethyl sulphate modification also supported this model.

scholarly journals High Prevalence of Growth Plate Gene Variants in Children With Familial Short Stature Treated With GH

2019 ◽  
Vol 104 (10) ◽  
pp. 4273-4281 ◽  
Author(s):  
Lukas Plachy ◽  
Veronika Strakova ◽  
Lenka Elblova ◽  
Barbora Obermannova ◽  
Stanislava Kolouskova ◽  
...  
Keyword(s):  
Short Stature ◽  
Growth Plate ◽  
Single Gene ◽  
Gene Variants ◽  
Genetic Etiology ◽  
Associated Proteins ◽  
Genetic Mechanisms ◽  
Standards And Guidelines ◽  
Minimum Height ◽  
Severe Short Stature

AbstractContextFamilial short stature (FSS) is a term describing a growth disorder that is vertically transmitted. Milder forms may result from the combined effect of multiple genes; more severe short stature is suggestive of a monogenic condition. The etiology of most FSS cases has not been thoroughly elucidated to date.ObjectivesTo identify the genetic etiology of severe FSS in children treated with GH because of the diagnosis of small for gestational age or GH deficiency (SGA/GHD).Design, Settings, and PatientsOf 736 children treated with GH because of GHD/SGA, 33 with severe FSS (life-minimum height −2.5 SD or less in both the patient and shorter parent) were included in the study. The genetic etiology was known in 5 of 33 children prior to the study [ACAN (in 2], NF1, PTPN11, and SOS1). In the remaining 28 of 33, whole-exome sequencing was performed. The results were evaluated using American College of Medical Genetics and Genomics standards and guidelines.ResultsIn 30 of 33 children (90%), we found at least one variant with potential clinical significance in genes known to affect growth. A genetic cause was elucidated in 17 of 33 (52%). Of these children, variants in growth plate-related genes were found in 9 of 17 [COL2A1, COL11A1, and ACAN (all in 2), FLNB, FGFR3, and IGF1R], and IGF-associated proteins were affected in 2 of 17 (IGFALS and HMGA2). In the remaining 6 of 17, the discovered genetic mechanisms were miscellaneous (TRHR, MBTPS2, GHSR, NF1, PTPN11, and SOS1).ConclusionsSingle-gene variants are frequent among families with severe FSS, with variants affecting the growth plate being the most prevalent.

scholarly journals Alternatively spliced mRNA variants of chloroplast ascorbate peroxidase isoenzymes in spinach leaves

1999 ◽  
Vol 338 (1) ◽  
pp. 41-48 ◽  
Author(s):  
Kazuya YOSHIMURA ◽  
Yukinori YABUTA ◽  
Masahiro TAMOI ◽  
Takahiro ISHIKAWA ◽  
Shigeru SHIGEOKA
Keyword(s):  
Ascorbate Peroxidase ◽  
Differential Splicing ◽  
Gene Encoding ◽  
S1 Nuclease ◽  
C Terminus ◽  
Mrna Variants ◽  
Alternatively Spliced ◽  
Splicing Efficiency ◽  
Spinach Leaves

We have previously shown that stromal and thylakoid-bound ascorbate peroxidase (APX) isoenzymes of spinach chloroplasts arise from a common pre-mRNA by alternative splicing in the C-terminus of the isoenzymes [Ishikawa, Yoshimura, Tamoi, Takeda and Shigeoka (1997) Biochem. J. 328, 795–800]. To explore the production of mature, functional mRNA encoding chloroplast APX isoenzymes, reverse transcriptase-mediated PCR and S1 nuclease protection analysis were performed with poly(A)+ RNA or polysomal RNA from spinach leaves. As a result, four mRNA variants, one form of thylakoid-bound APX (tAPX-I) and three forms of stromal APX (sAPX-I, sAPX-II and sAPX-III), were identified. The sAPX-I and sAPX-III mRNA species were generated through the excision of intron 11; they encoded the previously identified sAPX protein. Interestingly, the sAPX-II mRNA was generated by the insertion of intron 11 between exons 11 and 12. The use of this insertional sequence was in frame with the coding sequence and would lead to the production of a novel isoenzyme containing a C-terminus in which a seven-residue sequence replaced the last residue of the previously identified sAPX. The recombinant novel enzyme expressed in Escherichia coli showed the same enzymic properties (except for molecular mass) as the recombinant sAPX from the previously identified sAPX-I mRNA, suggesting that the protein translated from the sAPX-II mRNA is functional as a soluble APX in vivo. The S1 nuclease protection analysis showed that the expression levels of mRNA variants for sAPX and tAPX isoenzymes are in nearly equal quantities throughout the spinach leaves grown under normal conditions. The present results demonstrate that the expression of chloroplast APX isoenzymes is regulated by a differential splicing efficiency that is dependent on the 3´-terminal processing of ApxII, the gene encoding the chloroplast APX isoenzymes.

scholarly journals Influence of Intron Length on Alternative Splicing of CD44

1998 ◽  
Vol 18 (10) ◽  
pp. 5930-5941 ◽  
Author(s):  
Martyn V. Bell ◽  
Alison E. Cowper ◽  
Marie-Paule Lefranc ◽  
John I. Bell ◽  
Gavin R. Screaton
Keyword(s):  
Alternative Splicing ◽  
Genomic Structure ◽  
Single Gene ◽  
Intron Length ◽  
Protein Isoforms ◽  
Major Influence ◽  
Eukaryotic Genes ◽  
Alternatively Spliced ◽  
Alternatively Spliced Exons

ABSTRACT Although the splicing of transcripts from most eukaryotic genes occurs in a constitutive fashion, some genes can undergo a process of alternative splicing. This is a genetically economical process which allows a single gene to give rise to several protein isoforms by the inclusion or exclusion of sequences into or from the mature mRNA. CD44 provides a unique example; more than 1,000 possible isoforms can be produced by the inclusion or exclusion of a central tandem array of 10 alternatively spliced exons. Certain alternatively spliced exons have been ascribed specific functions; however, independent regulation of the inclusion or skipping of each of these exons would clearly demand an extremely complex regulatory network. Such a network would involve the interaction of many exon-specific trans-acting factors with the pre-mRNA. Therefore, to assess whether the exons are indeed independently regulated, we have examined the alternative exon content of a large number of individual CD44 cDNA isoforms. This analysis shows that the downstream alternatively spliced exons are favored over those lying upstream and that alternative exons are often included in blocks rather than singly. Using a novel in vivo alternative splicing assay, we show that intron length has a major influence upon the alternative splicing of CD44. We propose a kinetic model in which short introns may overcome the poor recognition of alternatively spliced exons. These observations suggest that for CD44, intron length has been exploited in the evolution of the genomic structure to enable tissue-specific patterns of splicing to be maintained.

scholarly journals Functional Dissection of an Alternatively Spliced Herpesvirus Gene by Splice Site Mutagenesis

2016 ◽  
Vol 90 (9) ◽  
pp. 4626-4636 ◽  
Author(s):  
Tim Schommartz ◽  
Stefan Loroch ◽  
Malik Alawi ◽  
Adam Grundhoff ◽  
Albert Sickmann ◽  
...  
Keyword(s):  
Functional Analysis ◽  
Viral Replication ◽  
Splice Site ◽  
Gene Knockout ◽  
Single Gene ◽  
Viral Gene ◽  
Gene Products ◽  
Knockout Mutants ◽  
Alternatively Spliced ◽  
Site Mutagenesis

ABSTRACTHerpesviruses have large and complex DNA genomes. The largest among the herpesviruses, those of the cytomegaloviruses, include over 170 genes. Although most herpesvirus gene products are expressed from unspliced transcripts, a substantial number of viral transcripts are spliced. Some viral transcripts are subject to alternative splicing, which leads to the expression of several proteins from a single gene. Functional analysis of individual proteins derived from an alternatively spliced gene is difficult, as deletion and nonsense mutagenesis, both common methods used in the generation of viral gene knockout mutants, affect several or all gene products at the same time. Here, we show that individual gene products of an alternatively spliced herpesvirus gene can be inactivated selectively by mutagenesis of the splice donor or acceptor site and by intron deletion or substitution mutagenesis. We used this strategy to dissect the essential M112/113 gene of murine cytomegalovirus (MCMV), which encodes the MCMV Early 1 (E1) proteins. The expression of each of the four E1 protein isoforms was inactivated individually, and the requirement for each isoform in MCMV replication was analyzed in fibroblasts, endothelial cells, and macrophages. We show that the E1 p87 isoform, but not the p33, p36, and p38 isoforms, is essential for viral replication in cell culture. Moreover, the presence of one of the two medium-size isoforms (p36 or p38) and the presence of intron 1, but not its specific sequence, are required for viral replication. This study demonstrates the usefulness of splice site mutagenesis for the functional analysis of alternatively spliced herpesvirus genes.IMPORTANCEHerpesviruses include up to 170 genes in their DNA genomes. The functions of most viral gene products remain poorly defined. The construction of viral gene knockout mutants has thus been an important tool for functional analysis of viral proteins. However, this strategy is of limited use when viral gene transcripts are alternatively spliced, leading to the expression of several proteins from a single gene. In this study, we showed, as a proof of principle, that each protein product of an alternatively spliced gene can be eliminated individually by splice site mutagenesis. Mutant viruses lacking individual protein products displayed different phenotypes, demonstrating that the products of alternatively spliced genes have nonredundant functions.

Alternative splicing can lead to chaos

2015 ◽  
Vol 13 (01) ◽  
pp. 1540003 ◽  
Author(s):  
Vitaly A. Likhoshvai ◽  
Vladislav V. Kogai ◽  
Stanislav I. Fadeev ◽  
Tamara M. Khlebodarova
Keyword(s):  
Alternative Splicing ◽  
Chaotic Dynamics ◽  
Single Gene ◽  
Regulatory Protein ◽  
Genetic System ◽  
Feedback Mechanism ◽  
Regulatory Proteins ◽  
Gene Encoding ◽  
Regulation Type ◽  
Alternatively Spliced

Alternative splicing is a widespread phenomenon in higher eukaryotes, where it serves as a mechanism to increase the functional diversity of proteins. This phenomenon has been described for different classes of proteins, including transcription regulatory proteins. We demonstrated that in the simplest genetic system model the formation of the alternatively spliced isoforms with opposite functions (activators and repressors) could be a cause of transition to chaotic dynamics. Under the simplest genetic system we understand a system consisting of a single gene encoding the structure of a transcription regulatory protein whose expression is regulated by a feedback mechanism. As demonstrated by numerical analysis of the models, if the synthesized isoforms regulate the expression of their own gene acting through different sites and independently of each other, for the generation of chaotic dynamics it is sufficient that the regulatory proteins have a dimeric structure. If regulatory proteins act through one site, the chaotic dynamics is generated in the system only when the repressor protein is either a tetrameric or a higher-dimensional multimer. In this case the activator can be a dimer. It was also demonstrated that if the transcription factor isoforms exhibit either activating or inhibiting activity and are lower-dimensional multimers (< 4), independently of the regulation type the model demonstrates either cyclic or stationary trajectories.

scholarly journals Characterization of CKβ8 and CKβ8-1: Two Alternatively Spliced Forms of Human β-Chemokine, Chemoattractants for Neutrophils, Monocytes, and Lymphocytes, and Potent Agonists at CC Chemokine Receptor 1

Blood ◽  
1998 ◽  
Vol 91 (9) ◽  
pp. 3118-3126 ◽  
Author(s):  
Byung-S. Youn ◽  
Shang M. Zhang ◽  
Hal E. Broxmeyer ◽  
Scott Cooper ◽  
Kathleen Antol ◽  
...  
Keyword(s):  
Amino Acids ◽  
Bone Marrow ◽  
Single Gene ◽  
The Other ◽  
New Members ◽  
Molecular Weights ◽  
Splicing Variant ◽  
Alternatively Spliced ◽  
Monocytes And Lymphocytes

Two new members of human β-chemokine cDNA were isolated based on structural and functional similarities to human leukotactin-1. One of these clones was identical to the previously isolated human β-chemokine, CKβ8, whereas the other is a splicing variant of CKβ8, therefore named CKβ8-1. CKβ8 was short in 51 nucleotides (17 amino acids) compared with CKβ8-1. The mature proteins of CKβ8-1 and CKβ8 consisted of 116 and 99 amino acids with calculated molecular weights of 12,500 and 10,950, respectively. Both CKβ8-1 and CKβ8 were potent agonists at CCR1. These chemokines chemoattracted neutrophils, monocytes, and lymphocytes. They also significantly suppressed colony formation by human bone marrow, granulocyte-macrophage, erythroid, and multipotential progenitor cells stimulated by combinations of growth factors. To our knowledge, this is the first example that an alternative splicing produces two active β-chemokines from a single gene.

scholarly journals 3D structure and in situ arrangements of CatSper channel in the sperm flagellum

2021 ◽  
Author(s):  
Yanhe Zhao ◽  
Huafeng Wang ◽  
Caroline Wiesehoefer ◽  
Naman B Shah ◽  
Evan Reetz ◽  
...  
Keyword(s):  
Electron Tomography ◽  
3D Structure ◽  
Human Sperm ◽  
Structural Basis ◽  
Sperm Flagellum ◽  
Mammalian Sperm ◽  
Zigzag Pattern ◽  
Associated Proteins

The sperm calcium channel CatSper plays a central role in successful fertilization as a primary Ca2+ gateway into the sperm flagellum. However, the complex subunit composition of CatSper has impeded its reconstitution in vitro and structural elucidation. Here, we applied cryo-electron tomography to visualize the macromolecular organization of the native CatSper channel complex in intact mammalian sperm, as well as identified three additional CatSper-associated proteins. The repeating CatSper units form long zigzag-rows in four nanodomains along the flagella. In both mouse and human sperm, each CatSper repeat consists of a tetrameric pore complex. Murine CatSper contains an additional outwardly directed wing-structure connected to the tetrameric channel. The majority of the extracellular domains form a canopy above each pore-forming channel that interconnects to a zigzag-shaped roof. The intracellular domains link two neighboring channel complexes to a diagonal array. The loss of this intracellular link in Efcab9-/- sperm distorts the longitudinally aligned zigzag pattern and compromises flagellar movement. This work offers unique insights into the mechanisms underlying the assembly and transport of the CatSper complex to generate the nanodomains and provides a long-sought structural basis for understanding CatSper function in the regulation of sperm motility.

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