An update on carbonic anhydrase-related proteins VIII, X and XI

Figures & data

Table 1.  Molecular properties of human and mouse CARPs.

	CARP isoforms	mRNA (kb)	Amino acid residues	Expected MW (kDa)	Chromosome localization
CARP VIII	Human	Multiple^b (2.4)	290	33	8q11-q12
	Mouse	Multiple^b (2.4)	291	33.1^c	—
CARP X	Human	2.8	328	37.6	17q24
	Mouse	2.6/3.0	328	37.6	—
CARP XI	Human	1.5	328	36.2	19q13.3
	Mouse	1.5	328	36.1	—

CARP, carbonic anhydrase-related protein.

^aThe longest cDNA clone deposited in GenBank is shown with the accession numbers. ^bAmong the five transcripts of different sizes, the strongest signal was observed at 2.4 kb. ^cMolecular size demonstrated by SDS-PAGE is 37.5 kDa³¹. Source: Adapted from Nishimori et al.³²

Table 2.  Distribution of CARP VIII protein and its mRNA in brain and other tissues.

	Adult human CNS^a		Human Fetal brain^b		Other tissues^c
	Regional distribution	Expression	Days of gestation	Expression	Human tissues	Expression
Cerebellum	Cortex	++	84	+	Liver	+
	Medulla	+	95	+	Lung	+
	Hippocampus	++	121	+	Hear	+
	Basal ganglia	++	141	+	Gut	+
			222	+	Thymus	+
Diencephalon	Thalamus	++			Kidney	+
	Substantia nigra	++
					Mouse tissues^d
					Cerebellum	+++
Cerebellum	Cortex, molecular layer	++			Frontal cortex	+
	Cortex, Purkinje cells	++			Parietal cortex	+
	Cortex, granular layer	–			Midbrain	+
	Medulla, dentate	+w			Eye	+
					Liver	++
Pons	Vestibular nuclei	++			Stomach	+w
	Abducens nucleus	++			Duodenum	+
	Pontine nuclei	++			Ileum	+
					Jejunum	+
Medulla	Olivary nuclei	++			Colon	+
	Others	++			Pancreas	–
					Spleen	+
Choroid plexus		++			Kidney	+
Pia arachnoid		++			Ovary	+w
					Heart	+
					Lung	+++
					Muscle	+w
					Testis	+w
					Uterus	–

Scores of expression level: +++, very strong expression, ++, strong expression was observed in most cells; +, moderate expression was observed in most cells; +w, weak expression was observed; –, no significant expression was observed.

CARP, carbonic anhydrase-related protein; CNS, central nervous system; RT-PCR, reverse transcription-PCR.

^a,bExpression analysis was done by immunohistochemistry using monoclonal antibodies against human CARPs⁵. ^cExpression of mRNA in mouse tissues using by northern hybridization method and RT-PCR^3,14. ^dExpression of mRNA was studied using quantitative PCR¹².

Figure 1.  Expression pattern of CA8 gene in human analyzed by microarray method. The expression profile figure is adapted from BioGPS (http://biogps.gnf.org, accessed July 2012). The figures shows expression of CA8 gene in different human tissues samples and expression is significantly high in cerebellum compared to other tissues²⁰.

Figure 2.  Expression of CA8 gene in normal and pathological conditions in human. Microarray data of the mRNA expression levels of human genes in normal and pathological tissues from MediSapiens. (http://www.medisapiens.com, accessed February 2012)²¹.

Figure 3.  Reduction in cerebellar volume of Saudi Arabian patients with a mutation in CA8 gene (Gln162-Arg). The brain magnetic resonance imaging (MRI) of the (A–D) patient 1, (E–H) patient 2, and the (I–L) patient 3. Figures show the progressive cerebellar loss in patient 1 between the sagittal T1 (A and C) and axial flair images (B and D) in the scans performed at 4 (A and B) and 8 years of age (C, D), respectively. Sagittal T1 images (E and I) show reduction in cerebellar volume more prominent in patient 3 (I) than in patient 2 (E). The volume loss is not associated with parenchymal signal abnormality as evident in images F, G, J, and K. Small sella turcica and skull base fatty replacement are also shown (E and I). Axial flair images (H and L) demonstrate deep periventricular white matter abnormalities with more involvement seen in patient 3 (L). Modified from Kaya et al.⁷

Figure 4.  Locations of the CA8 gene variation in members of Iraqi and Saudi Arabian family. (A) The location of the variation (yellow arrow, Glycine 162 Arginine) on hypothetical 3-D structure of carbonic anhydrase-related protein VIII (CARP VIII) protein as shown with incorrect numbering in Kaya et al.⁷ (B) The 3-D structure of human CARP VIII (PDB ID: 2W2J) showing the correct places of variations in the CARP VIII protein (substitution of amino acids) found in members of Iraqi (top right black arrow, Serine 100 Proline) and members of Saudi Arabian family (bottom right black arrow Glycine 162 Arginine), top left black arrow, the position of panel A correctly identified as Glycine 193.

Table 3.  Distribution of CARP X protein and its mRNA in brain and other tissues.

	Adult human CNS^a		Human fetal brain^b		Other tissues^c
	Regional distribution	Expression	Days of gestation	Expression	Human tissues	Expression
Cerebellum	Cortex	+w	84	+	Liver	+
	Medulla	++	95	+	Lung	+
	Hippocampus	–	121	+	Heart	+
	Basal ganglia	–	141	+	Gut	+
			222	+	Thymus	+
Diencephalon	Thalamus	–			Kidney	+
	Substantia nigra	–
					Mouse tissues^d
					Cerebellum	+++
Cerebellum	Cortex, molecular layer	–			Frontal cortex	++
	Cortex, Purkinje cells	+w			Parietal cortex	+++
	Cortex, granular layer	–			Midbrain	+
	Medulla, dentate	–			Eye	+w
					Liver	–
Pons	Vestibular nuclei	–			Stomach	–
	Abducens nucleus	–			Duodenum	–
	Pontine nuclei	–			Ilium	–
					Jejunum	–
Medulla	Olivary nuclei	+w			Colon	–
	Others	+w			Pancreas	–
					Spleen	–
Choroid plexus		+w			Kidney	–
Pia arachnoid		–			Ovary	–
					Heart	–
					Lung	–
					Muscle	–
					Testis	–
					Uterus	–

Immunohistochemistry, northern blot and RT-PCR score: ++, strong expression was observed in most Immunohistochemistry and RT-PCR score ++, strong expression was observed in most cells; +, moderate expression was observed in most cells; +w, weak expression was observed; –, no significant expression was observed.

CARP, carbonic anhydrase-related protein; CNS, central nervous system; RT-PCR, reverse transcription-PCR.

^a,bExpression analysis was done by immunohistochemistry using monoclonal antibodies against human CARPs⁵. ^cExpression was studied in human tissues using RT-PCR and northern blot method²³. ^dExpression of mRNA was studied using quantitative PCR¹².

Figure 5.  Expression profile of Car10 mRNA in murine tissues. The relative expression pattern of Car8 mRNAs in 20 different mouse tissues using RT-qPCR. The predominant expression of Car8 mRNA is seen central nervous system (CNS), expression being very high in cerebellum followed by parietal cortex and frontal cortex, very low level of expression was also seen in midbrain and eye (Table 3). Adopted from Aspatwar et al.¹²

Figure 6.  Immunohistochemical staining of carbonic anhydrase-related protein X (CARP X) protein in mouse tissues. Panel I. Expression of CARP X in the brain. (A) There was no expression in cerebellum. (B) The arrowheads in figure show CARP X-positive microcapillaries in the cerebrum. Panel II. Moderate CARP X expression was observed in the respiratory epithelium (arrowhead) of the lung (G). Moderate expression was also present in the gastric glands (arrows in panel D). The heart muscle cells occasionally showed extremely weak signals (I). The other tissues including the (A) liver, (B) pancreas, (C) submandibular gland, (E) colon, (F) small intestine, (H) skeletal muscle remained negative. Original magnifications are at ×20. Adopted from Aspatwar et al.¹²

Figure 7.  Expression pattern of CA10 gene in healthy human tissues analyzed by microarray method. The expression profile figure is adapted from BioGPS (http://biogps.gnf.org, accessed July 2012). The figures shows expression of CA10 gene in different human tissues samples and expression is markedly high in the pineal gland during the night compared to day time. Some expression can also be seen in cerebellum and other parts of the brain.²⁰

Figure 8.  Expression of CA10 gene in normal and pathological conditions in human: Microarray data of the mRNA expression levels of human genes in normal and pathological tissues from MediSapiens. (http://www.medisapiens.com, accessed February 2012)²¹.

Table 4.  Distribution of CARP XI protein and its mRNA in brain and other tissues.

	Adult human CNS^a		Human fetal brain^b		Other tissues^c
	Regional distribution	Expression	Days of gestation	Expression	Human tissues	Expression
Cerebrum	Cortex	+	84	+	Pancreas	++
	Medulla	–	95	+	Kidney	++
	Hippocampus	+	121	+	Liver	++
	Basal ganglia	+w	141	+	Salivary gland	++
			222	+	Spinal cord	++
Diencephalon	Thalamus	+			Thyroid	++
	Substantia nigra	–			Skeletal muscle+
					Ovary	+
					Small intestine	+
Cerebellum	Cortex, molecular layer	–			Colon	+
	Cortex, Purkinje cells	+			Peripheral–blood	+
	Cortex, granular layer	–			Leucocytes	+
	Medulla, dentate	+w			Stomach	+
					Lymphnode	+
Pons	Vestibular nuclei	+			Adrenal gland	+
	Abducens nucleus	+
	Pontine nuclei	++			Mouse-tissues^d
Medulla	Olivary nuclie	–			Cerebellum	++
	Others	–			Frontal cortex	+++
					Parietal cortex	+++
Choroid plexus		++			Midbrain	+
Pia arachnoid		++			Eye	+w
					Liver	–
					Stomach	–
					Duodenum	–
					Ilium	+w
					Jejunum	–
					Colon	+
					Pancreas	–
					Spleen	–
					Kidney	+
					Ovary	+
					Heart	+
					Lung	+
					Muscle	–
					Testis	–
					Uterus	–

Immunohistochemistry, northern blot and RT-PCR score: ++, strong expression was observed in most cells; +, moderate expression was observed in most cells; +w, weak expression was observed; –, no significant expression was observed.

CARP, carbonic anhydrase-related protein; CNS, central nervous system; RT-PCR, reverse transcription-PCR.

^a,bExpression analysis was done by immunohistochemistry using monoclonal antibodies against human CARPs⁵. ^cExpression of mRNA in human tissues by using Northern Blot and RT-PCR^28,30. ^dExpression of mRNA was studied using quantitative PCR¹².

Figure 9.  Distribution of Car11 mRNA in murine tissues. The relative expression pattern of Car11 mRNAs in 20 different mouse tissues analyzed using reverse transcription-quantitative PCR (RT-qPCR). The overall pattern of Car11 mRNA expression was similar to the expression of Car8 mRNA (Tables 2 and 4). The expression of mRNA and can be seen in most of the tissues. The Car11 mRNA was predominantly expressed in frontal cortex followed by significantly high level of expression in parietal cortex, cerebellum and midbrain. Low amount of Car11 mRNA signal was also seen in colon, kidney, ovary lung and eye (Table 4). Adopted from Aspatwar et al.¹²

Figure 10.  Immunohistochemical staining of carbonic anhydrase-related protein XI (CARP XI) protein in mouse tissues. Panel I. (A) Moderate expression of CARP XI in cerebellar Purkinje cells. (B) There was faint signal for CARP XI in cerebrum. Panel II. Weak immunoreactions for CARP XI in the crypts of Lieberkuhn (arrows in D), gastric glands (arrows in C), and renal tubule cells (F). Extremely weak signals can also be observed in the (A) liver, (E) colon and (I) testis. The (B) pancreas, (G) lung, and (H) skeletal muscle were all negative. Original magnifications are at ×20. Adopted from Aspatwar et al.¹²

Figure 11.  Expression pattern of CA11 gene in healthy human tissues analyzed by microarray method. The expression profile figure is adapted from BioGPS (http://biogps.gnf.org, accessed July 2012). The figures shows expression of CA11 gene in different human tissues samples and significant level of expression can only be seen in central nervous system²⁰.

Figure 12.  Expression of CA11 gene in normal and pathological conditions in human: Microarray data of the mRNA expression levels of human genes in normal and pathological tissues from MediSapiens. (http://www.medisapiens.com, accessed February 2012)²¹.

Bergenhem NC, Hallberg M, Wisén S. Molecular characterization of the human carbonic anhydrase-related protein (HCA-RP VIII). Biochim Biophys Acta 1998;1384:294–298.

 PubMed Web of Science ®Google Scholar

Supuran C, Scozzafava A, Conway, J, ed. Carbonic anhydrase: its inhibitors and activators. Washington, DC: CRC Press, 2004.

 Google Scholar

Taniuchi K, Nishimori I, Takeuchi T, Fujikawa-Adachi K, Ohtsuki Y, Onishi S. Developmental expression of carbonic anhydrase-related proteins VIII, X, and XI in the human brain. Neuroscience 2002a;112:93–99.

 PubMed Web of Science ®Google Scholar

Hirota J, Ando H, Hamada K, Mikoshiba K. Carbonic anhydrase-related protein is a novel binding protein for inositol 1,4,5-trisphosphate receptor type 1. Biochem J 2003;372:435–441.

 PubMedGoogle Scholar

Akisawa Y, Nishimori I, Taniuchi K, Okamoto N, Takeuchi T, Sonobe H et al. Expression of carbonic anhydrase-related protein CA-RP VIII in non-small cell lung cancer. Virchows Arch 2003;442:66–70.

 PubMed Web of Science ®Google Scholar

Aspatwar A, Tolvanen ME, Parkkila S. Phylogeny and expression of carbonic anhydrase-related proteins. BMC Mol Biol 2010;11:25.

 PubMedGoogle Scholar

Wu C, Orozco C, Boyer J, Leglise M, Goodale J, Batalov S et al. BioGPS: an extensible and customizable portal for querying and organizing gene annotation resources. Genome Biol 2009;10:R130.

 PubMed Web of Science ®Google Scholar

Kilpinen S, Autio R, Ojala K, Iljin K, Bucher E, Sara H et al. Systematic bioinformatic analysis of expression levels of 17,330 human genes across 9,783 samples from 175 types of healthy and pathological tissues. Genome Biol 2008;9:R139.

 PubMed Web of Science ®Google Scholar

Kaya N, Aldhalaan H, Al-Younes B, Colak D, Shuaib T, Al-Mohaileb F et al. Phenotypical spectrum of cerebellar ataxia associated with a novel mutation in the CA8 gene, encoding carbonic anhydrase (CA) VIII. Am J Med Genet B Neuropsychiatr Genet 2011;156B:826–834.

 PubMed Web of Science ®Google Scholar

Okamoto N, Fujikawa-Adachi K, Nishimori I, Taniuchi K, Onishi S. cDNA sequence of human carbonic anhydrase-related protein, CA-RP X: mRNA expressions of CA-RP X and XI in human brain. Biochim Biophys Acta 2001;1518:311–316.

 PubMedGoogle Scholar

Bellingham J, Gregory-Evans K, Gregory-Evans CY. Sequence and tissue expression of a novel human carbonic anhydrase-related protein, CARP-2, mapping to chromosome 19q13.3. Biochem Biophys Res Commun 1998;253:364–367.

 PubMed Web of Science ®Google Scholar

Fujikawa-Adachi K, Nishimori I, Taguchi T, Yuri K, Onishi S. cDNA sequence, mRNA expression, and chromosomal localization of human carbonic anhydrase-related protein, CA-RP XI. Biochim Biophys Acta 1999;1431:518–524.

 PubMedGoogle Scholar