Enzymatic amplification and molecular cloning of cDNA encoding the small and large subunits of bovine interleukin 12

Zarlenga, Dante S.; Canals, A.; Aschenbrenner, R.A.; Gasbarre, Louis C.

doi:10.1016/0925-4439(95)00042-3

Cited by 31 publications

(15 citation statements)

References 6 publications

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“…Plasmids used in this study were constructed using pCAT2AGUS, which was generated as previously described (15) and from the cloned cDNA encoding p40 and p35 using the sequence information provided by Zarlenga et al (16) The p40 subunit was amplified in the polymerase chain reaction (PCR) using the oligonucleotides p40.1 (59 -CCCCCCGTCGACGAT-GCACCCTCAGCAGTT-3 9 ), which introduced a SalI restriction site (underlined) immediately prior to the start codon (in bold) of the p40 leader sequence, and p40.2 (59 -CCCCCCG-CATGCACTGCAGGACACAGATGC-3 9 ), which removed the stop codon as well as introducing a SphI restriction site immediately following the p40 coding region. The PCR product was inserted into the SalI and SphI sites of pCAT2AGUS, to substitute the chloramphenicol acetyl transferase reporter gene (CAT), generating the construct p402AGUS.…”

Section: Generation Of Plasmid Constructsmentioning

confidence: 99%

Production of Interleukin-12 as a Self-Processing 2A Polypeptide

Chaplin¹,

Camon²,

Villarreal-Ramos³

et al. 1999

Journal of Interferon & Cytokine Research

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Interleukin-12 (IL-12) is a heterodimeric cytokine composed of two disulfide-linked subunits (p40 and p35) encoded by separate genes. We used the apparent autocleavage property of a 2A peptide from the foot-and-mouth disease virus (FMDV) to express bovine (Bo) IL-12 as a self-processing polypeptide (p402Ap35). We demonstrate that 2A will mediate the cleavage of p402Ap35 into two separate subunits in a manner similar to that observed during the processing of the FMDV polypeptide. Furthermore, this 2A polypeptide encoded a functional heterodimer, which elicited activities associated with IL-12 in other species. We propose that this strategy of self-processing polypeptides may be used in many applications where the coordinated and stoichiometric expression of complex proteins is required.

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Section: Generation Of Plasmid Constructsmentioning

confidence: 99%

Production of Interleukin-12 as a Self-Processing 2A Polypeptide

Chaplin¹,

Camon²,

Villarreal-Ramos³

et al. 1999

Journal of Interferon & Cytokine Research

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“…The bovine IL-12 molecule is structurally similar to that of humans and mice, and it increases IFN-␥ secretion in vitro by cattle peripheral blood mononuclear cells in response to viral antigens (57). IL-4 plays a role in protective immunity in cattle against helminth parasites (2).…”

mentioning

confidence: 99%

Pathogenesis and Immune Responses in Gnotobiotic Calves after Infection with the Genogroup II.4-HS66 Strain of Human Norovirus

Souza

Azevedo

Jung

et al. 2008

J Virol

151

131

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We previously characterized the pathogenesis of two host-specific bovine enteric caliciviruses (BEC), the GIII.2 norovirus (NoV) strain CV186-OH and the phylogenetically unassigned NB strain, in gnotobiotic (Gn) calves. In this study we evaluated the Gn calf as an alternative animal model to study the pathogenesis and host immune responses to the human norovirus (HuNoV) strain GII.4-HS66. The HuNoV HS66 strain caused diarrhea (five/five calves) and intestinal lesions (one/two calves tested) in the proximal small intestine (duodenum and jejunum) of Gn calves, with lesions similar to, but less severe than, those described for the Newbury agent 2 (NA-2) and NB BEC.

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“…To date bovine (15), cat (16 -18), dog (19,20), goat (J. C. Beyer, W. P. Cheevers, and N. M. Kumpula-McWhirter, unpublished observations), horse (21), pig (22), and sheep (23)(24)(25) homologues of IL12p40 and IL-12p35 subunits have been cloned. Some of them are currently being tested in clinical applications.…”

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confidence: 99%

Identification and Molecular Cloning of Functional Chicken IL-12

Degen

Daal

Zuilekom

et al. 2004

The Journal of Immunology

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By a combination of large-scale sequencing, bioinformatics, and traditional molecular biology, we identified the long-searched-for cDNA sequences encoding the homologues of the chicken IL-12p35 and IL-12p40 chains. These molecules are the first discovered nonmammalian IL-12 subunits. The homologies of the chicken IL-12p35 and IL-12p40 proteins to the corresponding known subunits of various species, i.e., humans, sheep, horse, cat, bovine, mouse, and woodchuck, ranged between 21 and 42%, respectively. The expression of IL-12 subunits was observed in lymphoid cells and proved to be dependent on the cell type and stimulus, while expression was not detected in stimulated primary chicken embryo fibroblast cells. Following transient expression of both molecules in COS-7 cells, we confirmed the necessity of heterodimerization into IL-12p70 to yield bioactivity as was also shown for its mammalian counterparts. The chicken IL-12p70 molecule, generated either by transient coexpression of monomeric IL-12p35 and monomeric IL-12p40 or as a fusion protein (as in a fusion linker construct), induced IFN-γ synthesis and proliferative activity of freshly exposed chicken splenocytes. The high degree of functional similarity between chicken IL-12 and IL-12 of higher mammalian vertebrates, despite their poor sequence homology, illustrates the conservation and vital importance of the IL-12 molecule since the evolutionary dichotomy of birds and mammals >300 million years ago. In this article, we describe the first nonmammalian IL-12 molecule and show that this chicken IL-12 molecule is bioactive.

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Enzymatic amplification and molecular cloning of cDNA encoding the small and large subunits of bovine interleukin 12

Cited by 31 publications

References 6 publications

Production of Interleukin-12 as a Self-Processing 2A Polypeptide

Production of Interleukin-12 as a Self-Processing 2A Polypeptide

Pathogenesis and Immune Responses in Gnotobiotic Calves after Infection with the Genogroup II.4-HS66 Strain of Human Norovirus

Identification and Molecular Cloning of Functional Chicken IL-12

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