Specific Interaction in Vitro and in Vivo of Glyceraldehyde-3-phosphate Dehydrogenase and LA Protein with Cis-acting RNAs of Human Parainfluenza Virus Type 3

De, Bishnu P.; Gupta, Sanhita; Zhao, Hong; Drazba, Judith A.; Banerjee, Amiya K.

doi:10.1074/jbc.271.40.24728

Cited by 108 publications

(94 citation statements)

References 38 publications

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“…Construction of two plasmids containing the 73-nt 3Ј genomic-sense (3Ј-GS) and leader sense (LS) RNAs, respectively, under control of the T7 promoter was described earlier (11). Radiolabeled RNAs were synthesized using these plasmid DNAs after linearization with HgaI in an in vitro transcription reaction mixture containing 500 M each ATP, GTP, and CTP; 25 M UTP; 50 Ci of [␣- 32 P]UTP; and 20 U of T7 RNA polymerase in a 50-l volume in accordance with the manufacturer's (Boehringer Mannheim) protocol.…”

Section: Methodsmentioning

confidence: 99%

“…In fact, comparison of the nucleotide sequences of different paramyxovirus 5Ј-and 3Ј-terminal RNA sequences identified highly conserved regions which may form the cognate site for binding of cellular proteins (50). In the case of HPIV3, we recently reported that cellular glyceraldehyde 3-phosphate dehydrogenase (GAPDH) and La protein interact with the viral cis-regulatory RNAs (11). Further analysis revealed that these cellular proteins are also packaged within the progeny virions, suggesting a role(s) for these cellular proteins in HPIV3 transcription and replication.…”

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confidence: 99%

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Specific Phosphorylated Forms of Glyceraldehyde 3-Phosphate Dehydrogenase Associate with Human Parainfluenza Virus Type 3 and Inhibit Viral Transcription In Vitro

Choudhary

Banerjee

2000

J Virol

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). To gain further insight into these molecular interactions, we analyzed the virion-associated GAPDH and La protein using two-dimensional gel electrophoresis and immunoblotting. The GAPDH was resolved into two major and one minor molecular species migrating in the pI range of 7.6 to 8.3, while the La protein was resolved into five molecular species in the pI range of 6.8 to 7.5. The GAPDH isoforms present in the virions were also detected in the cytoplasmic fraction of CV-1 cell extract, albeit as minor species. On the other hand, the multiple molecular forms of La protein as seen within the virions were readily detected in the total CV-1 cell extract. Further analysis of virion-associated GAPDH by in vivo labeling with [ 32 P]orthophosphate revealed the presence of multiple phosphorylated species. The phosphorylated species were able to bind specifically to the viral cis-acting 3 genome sense RNA but failed to bind to the leader sense RNA, as determined by gel mobility shift assay. In contrast, the La protein isoforms present within the virions were not phosphorylated and bound to the viral cis-acting RNAs in a phosphorylation-independent manner. The GAPDH isoforms purified from the CV-1 cell cytoplasmic fraction inhibited viral transcription in vitro. Consistent with this, flag-tagged recombinant GAPDH synthesized by using the vaccinia virus expression system also inhibited viral transcription. Together, these data indicate that specific phosphorylated forms of GAPDH associate with HPIV3 and are involved in the regulation of virus gene expression.Human parainfluenza virus type 3 (HPIV3), a paramyxovirus, is second only to respiratory syncytial virus in causing severe respiratory tract infections in children and young adults (6). The single-stranded RNA genome of HPIV3, 15,461 nucleotides (nt) long, is contained within a helical nucleocapsid (15). Three virus-encoded proteins, the nucleocapsid protein N (68 kDa), the phosphoprotein P (90 kDa), and the RNA polymerase L (257 kDa), are associated with the nucleocapsid to form a transcribing ribonucleoprotein (RNP) complex (1, 15). The N protein enwraps the genome RNA, while the L and P proteins together constitute the RNA-dependent RNA polymerase complex that transcribes and replicates the N-bound genome RNA. During transcription, the RNA polymerase synthesizes a short leader RNA, followed by six capped and polyadenylated mRNAs. During replication, on the other hand, a full-length plus-strand copy of the genome RNA is synthesized which, in turn, serves as the template for synthesis of the minus-strand genome RNA (1, 6, 15). Studies from our laboratory, as well as others, indicate that cellular cytoskeletal proteins such as actin and tubulin are involved in the activation of transcription of several paramyxoviruses (4,9,12,22,34,35). Similarly, cellular RNA binding proteins that interact with the viral cis-acting elements are believed to be involved in the regulation of transcription and replication (25,26,28).The minus-sense genomic 3Ј end and its complementary l...

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Section: Methodsmentioning

confidence: 99%

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confidence: 99%

Specific Phosphorylated Forms of Glyceraldehyde 3-Phosphate Dehydrogenase Associate with Human Parainfluenza Virus Type 3 and Inhibit Viral Transcription In Vitro

Choudhary

Banerjee

2000

J Virol

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“…It has previously been reported that GAPDH is localized in the perinuclear region and in polysomes in addition to its well-documented localization in the cytoplasm (De et al, 1996). Our investigations revealed that GAPDH is the major cytoplasmic (S-100) RNA-binding activity of BSC-1, HeLa and FRhK-4 cells, and that its association with ribosomal components (RSW) highly impaired this activity.…”

Section: Discussionmentioning

confidence: 62%

“…Specific binding of GAPDH to AU-rich elements (AREs) in the 39NTRs and association with polysomes has led to the suggestion that the protein may play a role in both regulation of ARE-dependent turnover and translation of mRNA (Nagy & Rigby, 1995). Furthermore, De et al (1996) demonstrated that GAPDH and La protein interact with two cis-acting RNAs of human parainfluenza virus type 3. Both RNAs contain AU-rich sequences and have the potential to form similar stem-loop structures.…”

Section: Discussionmentioning

confidence: 99%

Interaction of glyceraldehyde-3-phosphate dehydrogenase with secondary and tertiary RNA structural elements of the hepatitis A virus 3′ translated and non-translated regions

Dollenmaier

Weitz

2003

Journal of General Virology

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Proteins interacting with RNA structures at the 39 non-translated region (39NTR) of picornaviruses are probably important during viral RNA replication. We have shown previously that a dominant cellular cytoplasmic protein of 38 kDa (p38) interacts with the 39NTR and upstream regions of the hepatitis A virus (HAV) RNA (Kusov et al., J Virol 70, 1890-1897. Immunological and biochemical analyses of p38 have indicated that it is identical to GAPDH, which has previously been described as modulating translational regulation of the HAV RNA by interacting with the 59NTR (Schultz et al., J Biol Chem 271, 14134-14142, 1996). Three separate binding regions for GAPDH in the 39NTR and in the upstream 3D polymerase-coding region were identified. Structural analysis of these RNA regions by computer modelling and direct enzymatic cleavage suggested the presence of several AU-rich stem-loop structures having the potential for tertiary interactions. Binding of GAPDH to these structures was confirmed by RNA footprint analysis and resulted in the loss of double-stranded RNA regions. A different panel of RNA binding proteins (p28, p41 and p65) was detected in the ribosomal fractions of several cell lines (BSC-1, FRhK-4 and HeLa), whereas RNA binding of the GAPDH that was also present in these fractions was only marginal or absent.

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“…Of particular interest is the La autoantigen, which has been found to be associated with the 3ЈUTR (Ϫ) of Sindbis virus (32) and parainfluenza virus (10) in conjunction with glyceraldehyde-3-phosphate dehydrogenase. Some uncharacterized cellular proteins with molecular masses of 42 and 44 kDa, 36 and 38 kDa, and 108, 60, 50, and 42 kDa can stably bind to the 3ЈUTR (Ϫ) of Sindbis virus (31), poliovirus (35), and West Nile virus (38), respectively.…”

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confidence: 99%

Cellular Proteins from Human Monocytes Bind to Dengue 4 Virus Minus-Strand 3′ Untranslated Region RNA

Yocupicio-Monroy

Medina

Valle

et al. 2003

J Virol

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The synthesis of plus and minus RNA strands of several RNA viruses requires as a first step the interaction of some viral regulatory sequences with cellular and viral proteins. The dengue 4 virus genome, a singlestranded, positive-polarity RNA, is flanked by two untranslated regions (UTR) located in the 5 and 3 ends. The 3UTR in the minus-strand RNA [3UTR (؊)] has been thought to function as a promoter for the synthesis of plus-strand RNA. To study the initial interaction between this 3UTR and cellular and viral proteins, mobility shift assays were performed, and four ribonucleoprotein complexes (I through IV) were formed when uninfected and infected U937 cells (human monocyte cell line) interacted with the 3UTR (؊) of dengue 4 virus. Cross-linking assays with RNAs containing the complete 3UTR (؊) (nucleotides [nt] 101 to 1) or a partial sequence from nt 101 to 45 and nt 44 to 1 resulted in specific binding of some cellular proteins. Supermobility shift and immunoprecipitation assays demonstrated that the La protein forms part of these complexes. To determine the region in the 3 UTR that interacted with the La protein, two deletion mutants were generated. The mutant (del-96), with a deletion of nt 96 to 101, was unable to interact with the La protein, suggesting that La interacted with the 5 portion of the 3UTR (؊). Complex I, which was the main ribonucleoprotein complex formed with the 3UTR (؊) and which had the fastest electrophoretic migration, contained proteins such as calreticulin and protein disulfide isomerase, which constitute important components of the endoplasmic reticulum.Dengue virus (DEN), a mosquito-borne member of the Flaviviridae family, contains a single-stranded, positive-polarity RNA as a genome. The genomic RNA has a long open reading frame that encodes for a polyprotein that is processed during and after translation (reviewed in reference 8). This unique open reading frame is flanked by two untranslated regions located at the 5Ј and 3Ј ends (5ЈUTR and 3ЈUTR). In the 5Ј end, viral RNA contains a type I cap structure, and in the 3Ј end, it lacks the poly(A) track. The 5ЈUTR and 3ЈUTR of dengue 4 virus (DEN4) are 101 and 384 nucleotides (nt) in length, respectively. Both regions have been predicted to form stable stem-loop structures that are highly conserved among flaviviruses (5,7,26,37). The presence of conserved secondary structures within both UTRs suggests that they might contain cis-acting elements involved in translation and replication (17, 33). Positive-polarity RNA viruses use the positive-strand genomic 3ЈUTR [3ЈUTR (ϩ)] as a promoter for negativestrand RNA synthesis (Ϫ). In a similar manner, the 3ЈUTR present in the minus-polarity RNA could, like the 3ЈUTR (ϩ), contain promoter sequences for the transcription of positivepolarity RNAs. Besides, it has been revealed that DEN 5ЈUTR (ϩ) and 3ЈUTR (ϩ) contain several conserved elements which are involved in viral viability and replication (7,25,45). The presence of both regions is required for in vitro replication of DEN, supporting t...

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Specific Interaction in Vitro and in Vivo of Glyceraldehyde-3-phosphate Dehydrogenase and LA Protein with Cis-acting RNAs of Human Parainfluenza Virus Type 3

Cited by 108 publications

References 38 publications

Specific Phosphorylated Forms of Glyceraldehyde 3-Phosphate Dehydrogenase Associate with Human Parainfluenza Virus Type 3 and Inhibit Viral Transcription In Vitro

Specific Phosphorylated Forms of Glyceraldehyde 3-Phosphate Dehydrogenase Associate with Human Parainfluenza Virus Type 3 and Inhibit Viral Transcription In Vitro

Interaction of glyceraldehyde-3-phosphate dehydrogenase with secondary and tertiary RNA structural elements of the hepatitis A virus 3′ translated and non-translated regions

Cellular Proteins from Human Monocytes Bind to Dengue 4 Virus Minus-Strand 3′ Untranslated Region RNA

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