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Identification and preliminary characterization of mouse Adam33.

Gunn TM, Azarani A, Kim PH, Hyman RW, Davis RW, Barsh GS - BMC Genet. (2002)

Bottom Line: The metalloprotease-disintegrin family, or ADAM, proteins, are implicated in cell-cell interactions, cell fusion, and cell signaling, and are widely distributed among metazoan phyla.Orthologous relationships have been defined for a few ADAM proteins including ADAM10 (Kuzbanian), and ADAM17 (TACE), but evolutionary relationships are not clear for the majority of family members.Adam33 is expressed in the mouse adult brain and could play a role in complex processes that require cell-cell communication.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Pediatrics, and the Howard Hughes Medical Institute, Stanford University School of Medicine, Stanford, CA, USA.

ABSTRACT

Background: The metalloprotease-disintegrin family, or ADAM, proteins, are implicated in cell-cell interactions, cell fusion, and cell signaling, and are widely distributed among metazoan phyla. Orthologous relationships have been defined for a few ADAM proteins including ADAM10 (Kuzbanian), and ADAM17 (TACE), but evolutionary relationships are not clear for the majority of family members. Human ADAM33 refers to a testis cDNA clone that does not contain a complete open reading frame, but portions of the predicted protein are similar to Xenopus laevis ADAM13.

Results: In a 48 kb region of mouse DNA adjacent to the Attractin gene on mouse chromosome 2, we identified sequences very similar to human ADAM33. A full-length mouse cDNA was identified by a combination of gene prediction programs and RT-PCR, and the probable full-length human cDNA was identified by comparison to human genomic sequence in the homologous region on chromosome 20p13. Mouse ADAM33 is 44% identical to Xenopus laevis ADAM13, however a phylogenetic alignment and consideration of functional domains suggests that the two genes are not orthologous. Mouse Adam33 is widely expressed, most highly in the adult brain, heart, kidney, lung and testis.

Conclusions: While mouse ADAM33 is similar to Xenopus ADAM13 in sequence, further examination of its embryonic expression pattern, catalytic activity and protein interactions will be required to assess the functional relationship between these two proteins. Adam33 is expressed in the mouse adult brain and could play a role in complex processes that require cell-cell communication.

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Mouse Adam33 expression. (A) Northern hybridization assay of mouse Adam33 (using IMAGE clone 636599) on polyadenylated RNA from adult mouse tissues. (B) Expression profile of mouse Adam 33 in Origene Mouse Rapid-Scan cDNA panel. Panel consists of normalized cDNA from 24 mouse tissues or embryo stages, serially diluted over a 4-log range (1000X-1X), where the lowest concentration (IX) is approximately 1 pg. Adam33 expression was assayed by PCR amplification of a 368 bp fragment of the mouse Adam33 cDNA sequence (band shown). Lane 1: brain; 2: adrenal gland; 3: heart; 4: pancreas; 5: kidney; 6: uterus; 7: spleen; 8: prostate gland; 9: thymus; 10: day 8.5 embryo; 11: liver; 12: day 9.5 embryo; 13: stomach; 14: day 12.5 embryo; 15: small intestine; 16: day 19 embryo; 17: muscle; 18: virgin breast; 19: lung; 20: pregnant breast; 21: testis; 22: lactating breast; 23: skin; 24: involuting breast. Slight variation in the apparent mobility among lanes is due to sample differences; fragments of identical mobility among different tissue samples were produced when individual RT-PCR reactions were repeated. Absence of signal in the 1000× but presence in the 100× dilutions for two of the tissues are likely to be caused by contaminants in the RNA preparation that inhibited the RT-PCR reaction.
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Figure 3: Mouse Adam33 expression. (A) Northern hybridization assay of mouse Adam33 (using IMAGE clone 636599) on polyadenylated RNA from adult mouse tissues. (B) Expression profile of mouse Adam 33 in Origene Mouse Rapid-Scan cDNA panel. Panel consists of normalized cDNA from 24 mouse tissues or embryo stages, serially diluted over a 4-log range (1000X-1X), where the lowest concentration (IX) is approximately 1 pg. Adam33 expression was assayed by PCR amplification of a 368 bp fragment of the mouse Adam33 cDNA sequence (band shown). Lane 1: brain; 2: adrenal gland; 3: heart; 4: pancreas; 5: kidney; 6: uterus; 7: spleen; 8: prostate gland; 9: thymus; 10: day 8.5 embryo; 11: liver; 12: day 9.5 embryo; 13: stomach; 14: day 12.5 embryo; 15: small intestine; 16: day 19 embryo; 17: muscle; 18: virgin breast; 19: lung; 20: pregnant breast; 21: testis; 22: lactating breast; 23: skin; 24: involuting breast. Slight variation in the apparent mobility among lanes is due to sample differences; fragments of identical mobility among different tissue samples were produced when individual RT-PCR reactions were repeated. Absence of signal in the 1000× but presence in the 100× dilutions for two of the tissues are likely to be caused by contaminants in the RNA preparation that inhibited the RT-PCR reaction.

Mentions: Northern blot hybridization using a probe corresponding to the end of the Adam33 coding region and the beginning of the 3'UTR detects a 2.7 kb RNA in adult brain, heart, kidney, and lung, and a 2 kb RNA in testis (Figure 3A). The size of the 2.7 kb RNA corresponds exactly to the prediction based on sequence, but the molecular basis for the testis-specific 2 kb transcript is not known (attempts to amplify the cDNA from testis using primers in exon 1 and the 3'UTR were unsuccessful). Using a more sensitive assay for expression based on semi-quantitative RT-PCR revealed expression in all samples examined (Figure 3B), including total RNA from mouse embryos on days 8.5, 9.5, 12.5 and 19 of development (Figure 3B).


Identification and preliminary characterization of mouse Adam33.

Gunn TM, Azarani A, Kim PH, Hyman RW, Davis RW, Barsh GS - BMC Genet. (2002)

Mouse Adam33 expression. (A) Northern hybridization assay of mouse Adam33 (using IMAGE clone 636599) on polyadenylated RNA from adult mouse tissues. (B) Expression profile of mouse Adam 33 in Origene Mouse Rapid-Scan cDNA panel. Panel consists of normalized cDNA from 24 mouse tissues or embryo stages, serially diluted over a 4-log range (1000X-1X), where the lowest concentration (IX) is approximately 1 pg. Adam33 expression was assayed by PCR amplification of a 368 bp fragment of the mouse Adam33 cDNA sequence (band shown). Lane 1: brain; 2: adrenal gland; 3: heart; 4: pancreas; 5: kidney; 6: uterus; 7: spleen; 8: prostate gland; 9: thymus; 10: day 8.5 embryo; 11: liver; 12: day 9.5 embryo; 13: stomach; 14: day 12.5 embryo; 15: small intestine; 16: day 19 embryo; 17: muscle; 18: virgin breast; 19: lung; 20: pregnant breast; 21: testis; 22: lactating breast; 23: skin; 24: involuting breast. Slight variation in the apparent mobility among lanes is due to sample differences; fragments of identical mobility among different tissue samples were produced when individual RT-PCR reactions were repeated. Absence of signal in the 1000× but presence in the 100× dilutions for two of the tissues are likely to be caused by contaminants in the RNA preparation that inhibited the RT-PCR reaction.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC88886&req=5

Figure 3: Mouse Adam33 expression. (A) Northern hybridization assay of mouse Adam33 (using IMAGE clone 636599) on polyadenylated RNA from adult mouse tissues. (B) Expression profile of mouse Adam 33 in Origene Mouse Rapid-Scan cDNA panel. Panel consists of normalized cDNA from 24 mouse tissues or embryo stages, serially diluted over a 4-log range (1000X-1X), where the lowest concentration (IX) is approximately 1 pg. Adam33 expression was assayed by PCR amplification of a 368 bp fragment of the mouse Adam33 cDNA sequence (band shown). Lane 1: brain; 2: adrenal gland; 3: heart; 4: pancreas; 5: kidney; 6: uterus; 7: spleen; 8: prostate gland; 9: thymus; 10: day 8.5 embryo; 11: liver; 12: day 9.5 embryo; 13: stomach; 14: day 12.5 embryo; 15: small intestine; 16: day 19 embryo; 17: muscle; 18: virgin breast; 19: lung; 20: pregnant breast; 21: testis; 22: lactating breast; 23: skin; 24: involuting breast. Slight variation in the apparent mobility among lanes is due to sample differences; fragments of identical mobility among different tissue samples were produced when individual RT-PCR reactions were repeated. Absence of signal in the 1000× but presence in the 100× dilutions for two of the tissues are likely to be caused by contaminants in the RNA preparation that inhibited the RT-PCR reaction.
Mentions: Northern blot hybridization using a probe corresponding to the end of the Adam33 coding region and the beginning of the 3'UTR detects a 2.7 kb RNA in adult brain, heart, kidney, and lung, and a 2 kb RNA in testis (Figure 3A). The size of the 2.7 kb RNA corresponds exactly to the prediction based on sequence, but the molecular basis for the testis-specific 2 kb transcript is not known (attempts to amplify the cDNA from testis using primers in exon 1 and the 3'UTR were unsuccessful). Using a more sensitive assay for expression based on semi-quantitative RT-PCR revealed expression in all samples examined (Figure 3B), including total RNA from mouse embryos on days 8.5, 9.5, 12.5 and 19 of development (Figure 3B).

Bottom Line: The metalloprotease-disintegrin family, or ADAM, proteins, are implicated in cell-cell interactions, cell fusion, and cell signaling, and are widely distributed among metazoan phyla.Orthologous relationships have been defined for a few ADAM proteins including ADAM10 (Kuzbanian), and ADAM17 (TACE), but evolutionary relationships are not clear for the majority of family members.Adam33 is expressed in the mouse adult brain and could play a role in complex processes that require cell-cell communication.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Pediatrics, and the Howard Hughes Medical Institute, Stanford University School of Medicine, Stanford, CA, USA.

ABSTRACT

Background: The metalloprotease-disintegrin family, or ADAM, proteins, are implicated in cell-cell interactions, cell fusion, and cell signaling, and are widely distributed among metazoan phyla. Orthologous relationships have been defined for a few ADAM proteins including ADAM10 (Kuzbanian), and ADAM17 (TACE), but evolutionary relationships are not clear for the majority of family members. Human ADAM33 refers to a testis cDNA clone that does not contain a complete open reading frame, but portions of the predicted protein are similar to Xenopus laevis ADAM13.

Results: In a 48 kb region of mouse DNA adjacent to the Attractin gene on mouse chromosome 2, we identified sequences very similar to human ADAM33. A full-length mouse cDNA was identified by a combination of gene prediction programs and RT-PCR, and the probable full-length human cDNA was identified by comparison to human genomic sequence in the homologous region on chromosome 20p13. Mouse ADAM33 is 44% identical to Xenopus laevis ADAM13, however a phylogenetic alignment and consideration of functional domains suggests that the two genes are not orthologous. Mouse Adam33 is widely expressed, most highly in the adult brain, heart, kidney, lung and testis.

Conclusions: While mouse ADAM33 is similar to Xenopus ADAM13 in sequence, further examination of its embryonic expression pattern, catalytic activity and protein interactions will be required to assess the functional relationship between these two proteins. Adam33 is expressed in the mouse adult brain and could play a role in complex processes that require cell-cell communication.

Show MeSH
Related in: MedlinePlus