Carnitine transporter OCTN2 mutations in systemic primary carnitine deficiency: a novel Arg169Gln mutation and a recurrent Arg282ter mutation associated with an unconventional splicing abnormality.

Systemic primary carnitine deficiency (CDSP, MIM 212140) is a disorder of fatty acid oxidation manifesting in acute metabolic decompensation or in progressive cardiomyopathy and muscle weakness. Mutations in the plasmalemmal organic cation/carnitine transporter OCTN2 were recently identified in CDSP patients of diverse ethnic backgrounds. We have performed OCTN2 mutation analysis in two unrelated German patients with primary carnitine deficiency and identified three molecular abnormalities. On one of the four chromosomes analyzed, we detected an Arg169Gln missense mutation that affects an arginine residue absolutely conserved in the entire transporter superfamily to which OCTN2 belongs. On the three other chromosomes, we found an Arg282ter nonsense mutation in exon 5. This mutation is embedded into different haplotypes of closely spaced intragenic dimorphisms in our two patients and was recently described in a patient of Asiatic Indian background, so it appears to be a recurrent or ancient founder mutation that may account for more CDSP cases. Finally, we found that the Arg282ter nonsense mutation is associated with a splicing abnormality at the intron 6/exon 7 junction. However, no mutations are present in exon 6, intron 6, or exon 7, suggesting that defective splicing of exon 7 on the Arg282ter allele is due to an unconventional, long-distance mechanism.     

Mutations in novel organic cation transporter (OCTN2), an organic cation/carnitine transporter, with differential effects on the organic cation transport function and the carnitine transport function

Novel organic cation transporter (OCTN2) is an organic cation/carnitine transporter, and two missense mutations, L352R and P478L, in OCTN2 have been identified as the cause for primary carnitine deficiency. In the present study, we assessed the influence of these two mutations on the carnitine transport function and the organic cation transport function of OCTN2. The L352R mutation resulted in a complete loss of both transport functions. In contrast, the P478L mutation resulted in a complete loss of only the carnitine transport function but significantly stimulated the organic cation transport function. Studies with human OCTN2/rat OCTN2 chimeric transporters indicated that the carnitine transport site and the organic cation transport site were not identical. Because carnitine transport is Na(+)-dependent whereas organic cation transport is Na(+)-independent, we investigated the possibility that the P478L mutation affected Na(+) binding. The Na(+) activation kinetics were found to be similar for the P478L mutant and wild type OCTN2. We then mutated nine different tyrosine residues located in or near transmembrane domains and assessed the transport function of these mutants. One of these mutations, Y211F, was found to have differential influence on the two transport activities of OCTN2 as did the P478L mutation. However, the Na(+) activation kinetics were not affected. These findings are of clinical relevance to patients with primary carnitine deficiency because whereas each and every mutation in these patients is expected to result in the loss of the carnitine transport function, all of these mutations may not interfere with the organic cation transport function.     

Molecular and functional characterization of organic cation/carnitine transporter family in mice

Carnitine is essential for beta-oxidation of fatty acids, and a defect of cell membrane transport of carnitine leads to fatal systemic carnitine deficiency. We have already shown that a defect of the organic cation/carnitine transporter OCTN2 is a primary cause of systemic carnitine deficiency. In the present study, we further isolated and characterized new members of the OCTN family, OCTN1 and -3, in mice. All three members were expressed commonly in kidney, and OCTN1 and -2 were also expressed in various tissues, whereas OCTN3 was characterized by predominant expression in testis. When their cDNAs were transfected into HEK293 cells, the cells exhibited transport activity for carnitine and/or the organic cation tetraethylammonium (TEA). Carnitine transport by OCTN1 and OCTN2 was Na(+)-dependent, whereas that by OCTN3 was Na(+)-independent. TEA was transported by OCTN1 and OCTN2 but not by OCTN3. The relative uptake activity ratios of carnitine to TEA were 1.78, 11.3, and 746 for OCTN1, -2, and -3, respectively, suggesting high specificity of OCTN3 for carnitine and significantly lower carnitine transport activity of OCTN1. Thus, OCTN3 is unique in its limited tissue distribution and Na(+)-independent carnitine transport, whereas OCTN1 efficiently transported TEA with minimal expression of carnitine transport activity and may have a different role from other members of the OCTN family.     

beta-lactam antibiotics as substrates for OCTN2, an organic cation/carnitine transporter

Therapeutic use of cephaloridine, a beta-lactam antibiotic, in humans is associated with carnitine deficiency. A potential mechanism for the development of carnitine deficiency is competition between cephaloridine and carnitine for the renal reabsorptive process. OCTN2 is an organic cation/carnitine transporter that is responsible for Na(+)-coupled transport of carnitine in the kidney and other tissues. We investigated the interaction of several beta-lactam antibiotics with OCTN2 using human cell lines that express the transporter constitutively as well as using cloned human and rat OCTN2s expressed heterologously in human cell lines. The beta-lactam antibiotics cephaloridine, cefoselis, cefepime, and cefluprenam were found to inhibit OCTN2-mediated carnitine transport. These antibiotics possess a quaternary nitrogen as does carnitine. Several other beta-lactam antibiotics that do not possess this structural feature did not interact with OCTN2. The interaction of cephaloridine with OCTN2 is competitive with respect to carnitine. Interestingly, many of the beta-lactam antibiotics that were not recognized by OCTN2 were good substrates for the H(+)-coupled peptide transporters PEPT1 and PEPT2. In contrast, cephaloridine, cefoselis, cefepime, and cefluprenam, which were recognized by OCTN2, did not interact with PEPT1 and PEPT2. The interaction of cephaloridine with OCTN2 was Na(+)-dependent, whereas the interaction of cefoselis and cefepime with OCTN2 was largely Na(+)-independent. Furthermore, the Na(+)-dependent, OCTN2-mediated cellular uptake of cephaloridine could be demonstrated by direct uptake measurements. These studies show that OCTN2 plays a crucial role in the pharmacokinetics and therapeutic efficacy of certain beta-lactam antibiotics such as cephaloridine and that cephaloridine-induced carnitine deficiency is likely to be due to inhibition of carnitine reabsorption in the kidney.     

Abnormal sodium stimulation of carnitine transport in primary carnitine deficiency

Primary carnitine deficiency is an autosomal recessive disorder of fatty acid oxidation characterized by hypoketotic hypoglycemia and skeletal and cardiac myopathy. It is caused by mutations in the sodium-dependent carnitine cotransporter OCTN2. The majority of natural mutations identified in this and other Na(+)/solute symporters introduce premature termination codons or impair insertion of the mutant transporter on the plasma membrane. Here we report that a missense mutation (E452K) identified in one patient with primary carnitine deficiency did not affect membrane targeting, as assessed with confocal microscopy of transporters tagged with the green fluorescent protein, but reduced carnitine transport by impairing sodium stimulation of carnitine transport. The natural mutation increased the concentration of sodium required to half-maximally stimulate carnitine transport (K(Na)) from the physiological value of 11.6 to 187 mm. Substitution of Glu(452) with glutamine (E452Q), aspartate (E452D), or alanine (E452A) caused intermediate increases in the K(Na). Carnitine transport decreased exponentially with increased K(Na). The E452K mutation is the first natural mutation in a mammalian cotransporter affecting sodium-coupled solute transfer and identifies a novel domain of the OCTN2 cotransporter involved in transmembrane sodium/solute transfer.     

A double missense mutation in the ATM gene of a Dutch family with ataxia telangiectasia

Ataxia telangiectasia (AT) is an autosomal recessive disorder characterized by cerebellar ataxia, telangiectasia, immunodeficiency, elevated α-fetoprotein levels, chromosomal instability, predisposition to cancer, and radiation sensitivity. We report the identification of a new, double missense mutation in the ataxia telangiectasia gene (ATM) of a Dutch family. This homozygous mutation consists of two consecutive base substitutions in exon 55: a T→G transversion at position 7875 of the ATM cDNA and a G→C transversion at position 7876. These transversions were confirmed by polymerase chain reaction/primer-induced restriction analysis with CelII. The double base substitution results in an amino acid change of an aspartic acid to a glutamic acid at codon 2625 and of an alanine to a proline at codon 2626 of the ATM protein. Both amino acids are conserved between the ATM protein and its functional homolog, the Atm gene product in the mouse. Furthermore, the Chou-Fasman and Robson predictions both demonstrate a change in the secondary structure of the ATM protein carrying the D2625E/A2626P mutation. These findings suggest that the double base substitution in the ATM gene is a disease-causing mutation. Received: 6 October 1997 / Accepted: 5 November 1997     

Tyrosine residues affecting sodium stimulation of carnitine transport in the OCTN2 carnitine/organic cation transporter

Primary carnitine deficiency is a disorder of fatty acid oxidation caused by mutations in the Na+-dependent carnitine/organic cation transporter OCTN2. Studies with tyrosyl group-modifying reagents support the involvement of tyrosine residues in Na+ binding by sodium-coupled transporters. Here we report two new patients with carnitine deficiency caused by mutations affecting tyrosyl residues (Y447C and Y449D) close to a residue (Glu-452) previously shown to affect sodium stimulation of carnitine transport. Kinetic analysis indicated that the Y449D substitution, when expressed in Chinese hamster ovary cells, increased the concentration of sodium required to half-maximally stimulate carnitine transport from 14.8 +/- 1.8 to 34.9 +/- 5.8 mM (p<0.05), whereas Y447C completely abolished carnitine transport. Substitution of these tyrosine residues with phenylalanine restored normal carnitine transport in Y449F but resulted in markedly impaired carnitine transport by Y447F. This was associated with an increase in the concentration of sodium required to half-maximally stimulate carnitine transport to 57.8 +/- 7.4 mM (p<0.01 versus normal OCTN2). The Y447F and Y449D mutant transporters retained their ability to transport the organic cation tetraethylammonium indicating that their effect on carnitine transport was specific and likely associated with the impaired sodium stimulation of carnitine transport. By contrast, the Y447C natural mutation abolished the transport of organic cations in addition to carnitine. Confocal microscopy of OCTN2 transporters tagged with green fluorescent protein indicated that the Y447C mutant transporters failed to reach the plasma membrane, whereas Y447F, Y449D, and Y449F had normal membrane localization. These natural mutations identify tyrosine residues possibly involved in coupling the sodium electrochemical gradient to transmembrane solute transfer in the sodium-dependent co-transporter OCTN2.     

Inheritance of the S113L mutation within an inbred family with carnitine palmitoyltransferase enzyme deficiency

Deficiency of carnitine palmitoyltransferase type II (CPT II) is a clinically heterogeneous autosomal recessive disorder of lipid metabolism. The most common mutation in the CPT 11 gene is the S113L mutation, which substitutes leucine for serine at amino acid position 113. We studied an inbred family with three affected cousins with CPT II deficiency and found the S113L mutation to be present in a homozygous state in all three patients. Pedigree analysis traced the S 113L mutation back to one common ancestor. Although the patients in this family have an identical genotype at the CPT II locus, their clinical picture ranges from asymptomatic to lethal.     

Functional domains in the carnitine transporter OCTN2, defective in primary carnitine deficiency

Primary carnitine deficiency is an autosomal recessive disorder of fatty acid oxidation characterized by hypoketotic hypoglycemia and skeletal and cardiac myopathy. It is caused by mutations in the Na+-dependent organic cation transporter, OCTN2. To define the domains involved in carnitine recognition, we evaluated chimeric transporters created by swapping homologous domains between OCTN1, which does not transport carnitine, and OCTN2. Substitution of the C terminus of OCTN2 (amino acid residues 342-557) with the corresponding residues of OCTN1 completely abolished carnitine transport. The progressive substitution of the N terminus of OCTN2 with OCTN1 resulted in a decrease in carnitine transport associated with a progressive increase in the Km toward carnitine from 3.9 +/- 0.5 to 141 +/- 19 microM. The largest drop in carnitine transport (and increase in Km toward carnitine) was observed with the substitution of residues 341-454 of OCTN2. An additional chimeric transporter (CHIM-9) in which only residues 341-454 of OCTN2 were substituted by OCTN1 had markedly reduced carnitine transport, with an elevated Km toward carnitine (63 +/- 5 microM). Site-directed mutagenesis and introduction of residues nonconserved between OCTN1 and OCTN2 in the OCTN2 cDNA indicated that the R341A, L409W, L424Y, and T429I substitutions significantly decreased carnitine transport. Single substitutions did not increase the Km toward carnitine. By contrast, the combination of three of these substitutions (R341W + L409W + T429I) greatly decreased carnitine transport and increased the Km toward carnitine (20.2 +/- 4.5 microm). The Arg-341, Leu-409, and Thr-429 residues are all located in predicted transmembrane domains. Involvement of these residues in carnitine transport was further supported by the partial restoration of carnitine transport by the introduction of these OCTN2 residues in the OCTN1 portion of CHIM-9. These studies indicate that multiple domains of the OCTN2 transporter are required for carnitine transport and identify transmembrane residues important for carnitine recognition.     

OCTN2VT, a splice variant of OCTN2, does not transport carnitine because of the retention in the endoplasmic reticulum caused by insertion of 24 amino acids in the first extracellular loop of OCTN2

A novel organic cation transporter OCTN2 is indispensable for carnitine transport across plasma membrane and subsequent fatty acid metabolism in the mitochondria. Here, we report a novel splice variant of OCTN2 (OCTN2VT), in which a 72-base-pair sequence located in the first intron of OCTN2 gene was spliced between exons 1 and 2 of OCTN2, causing the insertion of 24 amino acids in the first extracellular loop of OCTN2. Despite the similarity between OCTN2 and OCTN2VT regarding primary structure and tissue distribution, their biochemical characteristics were significantly different. OCTN2 was expressed on the plasma membrane with robust N-glycosylation, whereas OCTN2VT was retained in the endoplasmic reticulum (ER) with poor N-glycosylation. In addition, the retention in the ER caused no carnitine uptake into the cells. These results demonstrate that the biochemical and functional characteristics of OCTN2VT are distinct from OCTN2 due to the insertion of 24 amino acids in the first extracellular loop.     

Wide tolerance to amino acids substitutions in the OCTN1 ergothioneine transporter

BackgroundOrganic cation transporters transfer solutes with a positive charge across the plasma membrane. The novel organic cation transporter 1 (OCTN1) and 2 (OCTN2) transport ergothioneine and carnitine, respectively. Mutations in the SLC22A5 gene encoding OCTN2 cause primary carnitine deficiency, a recessive disorders resulting in low carnitine levels and defective fatty acid oxidation. Variations in the SLC22A4 gene encoding OCTN1 are associated with rheumatoid arthritis and Crohn disease.MethodsHere we evaluate the functional properties of the OCTN1 transporter using chimeric transporters constructed by fusing different portion of the OCTN1 and OCTN2 cDNAs. Their relative abundance and subcellular distribution was evaluated through western blot analysis and confocal microscopy.ResultsSubstitutions of the C-terminal portion of OCTN1 with the correspondent residues of OCTN2 generated chimeric OCTN transporters more active than wild-type OCTN1 in transporting ergothioneine. Additional single amino acid substitutions introduced in chimeric OCTN transporters further increased ergothioneine transport activity. Kinetic analysis indicated that increased transport activity was due to an increased V_max, with modest changes in K_m toward ergothioneine.ConclusionsOur results indicate that the OCTN1 transporter is tolerant to extensive amino acid substitutions. This is in sharp contrast to the OCTN2 carnitine transporter that has been selected for high functional activity through evolution, with almost all substitutions reducing carnitine transport activity.General significanceThe widespread tolerance of OCTN1 to amino acid substitutions suggests that the corresponding SLC22A4 gene may have derived from a recent duplication of the SLC22A5 gene and might not yet have a defined physiological role.     

Functional relevance of carnitine transporter OCTN2 to brain distribution of l-carnitine and acetyl-l-carnitine across the blood–brain barrier

Transport of L-[3H]carnitine and acetyl-L-[3H]carnitine at the blood-brain barrier (BBB) was examined by using in vivo and in vitro models. In vivo brain uptake of acetyl-L-[3H]carnitine, determined by a rat brain perfusion technique, was decreased in the presence of unlabeled acetyl-L-carnitine and in the absence of sodium ions. Similar transport properties for L-[3H]carnitine and/or acetyl-L-[3H]carnitine were observed in primary cultured brain capillary endothelial cells (BCECs) of rat, mouse, human, porcine and bovine, and immortalized rat BCECs, RBEC1. Uptakes of L-[3H]carnitine and acetyl-L-[3H]carnitine by RBEC1 were sodium ion-dependent, saturable with K(m) values of 33.1 +/- 11.4 microM and 31.3 +/- 11.6 microM, respectively, and inhibited by carnitine analogs. These transport properties are consistent with those of carnitine transport by OCTN2. OCTN2 was confirmed to be expressed in rat and human BCECs by an RT-PCR method. Furthermore, the uptake of acetyl-L-[3H]carnitine by the BCECs of juvenile visceral steatosis (jvs) mouse, in which OCTN2 is functionally defective owing to a genetical missense mutation of one amino acid residue, was reduced. The brain distributions of L-[3H]carnitine and acetyl-L-[3H]carnitine in jvs mice were slightly lower than those of wild-type mice at 4 h after intravenous administration. These results suggest that OCTN2 is involved in transport of L-carnitine and acetyl-L-carnitine from the circulating blood to the brain across the BBB.     

Carnitine Deficiency in OCTN2?/? Newborn Mice Leads to a Severe Gut and Immune Phenotype with Widespread Atrophy,Apoptosis and a Pro-Inflammatory Response

We have investigated the gross, microscopic and molecular effects of carnitine deficiency in the neonatal gut using a mouse model with a loss-of-function mutation in the OCTN2 (SLC22A5) carnitine transporter. The tissue carnitine content of neonatal homozygous (OCTN2^−/−) mouse small intestine was markedly reduced; the intestine displayed signs of stunted villous growth, early signs of inflammation, lymphocytic and macrophage infiltration and villous structure breakdown. Mitochondrial β-oxidation was active throughout the GI tract in wild type newborn mice as seen by expression of 6 key enzymes involved in β-oxidation of fatty acids and genes for these 6 enzymes were up-regulated in OCTN2^−/− mice. There was increased apoptosis in gut samples from OCTN2^−/− mice. OCTN2^−/− mice developed a severe immune phenotype, where the thymus, spleen and lymph nodes became atrophied secondary to increased apoptosis. Carnitine deficiency led to increased expression of CD45-B220⁺ lymphocytes with increased production of basal and anti-CD3-stimulated pro-inflammatory cytokines in immune cells. Real-time PCR array analysis in OCTN2^−/− mouse gut epithelium demonstrated down-regulation of TGF-β/BMP pathway genes. We conclude that carnitine plays a major role in neonatal OCTN2^−/− mouse gut development and differentiation, and that severe carnitine deficiency leads to increased apoptosis of enterocytes, villous atrophy, inflammation and gut injury.     

GFP-Human high-affinity carnitine transporter OCTN2 protein: subcellular localization and functional restoration of carnitine uptake in mutant cell lines with the carnitine transporter defect.

Individuals with the plasmalemmal high-affinity carnitine transporter defect present with progressive infantile-onset carnitine-responsive cardiomyopathy, lipid storage myopathy, recurrent hypoglycemic hypoketotic encephalopathy, and failure to thrive. The carnitine uptake defect (CUD) has been documented in their cultured skin fibroblasts, lymphoblasts, and/or myoblasts. The cDNA encoding the high-affinity sodium-dependent human carnitine transporter OCTN2 has recently been cloned. We used the green fluorescent protein (GFP) as a living marker for positively transfected cells in our expression studies of the high-affinity carnitine transporter OCTN2 cDNA in cell lines with the CUD. Transfection of cell lines from 12 unrelated patients (nine fibroblast and three lymphoblastoid) with a GFP construct harboring the wild-type full-length OCTN2 cDNA was done using LipoTAXI. Transient and stable expression of the recombinant GFP-human carnitine transporter OCTN2 cDNA was surveyed, and transient transfection of the fibroblast and stable transfection of the lymphoblastoid cell lines were achieved. There was functional restoration of carnitine uptake in the transfected mutant cell lines, thereby confirming the identity of the transfected cDNA. In addition, we report the first demonstration of the subcellular localization of an in-frame fusion GFP-human high-affinity carnitine transporter OCTN2 protein in the plasma membrane by confocal laser-scanning fluorescence microscopy.     

Glycosylation of the OCTN2 carnitine transporter: Study of natural mutations identified in patients with primary carnitine deficiency

Primary carnitine deficiency is caused by impaired activity of the Na⁺-dependent OCTN2 carnitine/organic cation transporter. Carnitine is essential for entry of long-chain fatty acids into mitochondria and its deficiency impairs fatty acid oxidation. Most missense mutations identified in patients with primary carnitine deficiency affect putative transmembrane or intracellular domains of the transporter. Exceptions are the substitutions P46S and R83L located in an extracellular loop close to putative glycosylation sites (N57, N64, and N91) of OCTN2. P46S and R83L impaired glycosylation and maturation of OCTN2 transporters to the plasma membrane. We tested whether glycosylation was essential for the maturation of OCTN2 transporters to the plasma membrane. Substitution of each of the three asparagine (N) glycosylation sites with glutamine (Q) decreased carnitine transport. Substitution of two sites at a time caused a further decline in carnitine transport that was fully abolished when all three glycosylation sites were substituted by glutamine (N57Q/N64Q/N91Q). Kinetic analysis of carnitine and sodium-stimulated carnitine transport indicated that all substitutions decreased the Vmax for carnitine transport, but N64Q/N91Q also significantly increased the Km toward carnitine, indicating that these two substitutions affected regions of the transporter important for substrate recognition. Western blot analysis confirmed increased mobility of OCTN2 transporters with progressive substitutions of asparagines 57, 64 and/or 91 with glutamine. Confocal microscopy indicated that glutamine substitutions caused progressive retention of OCTN2 transporters in the cytoplasm, up to full retention (such as that observed with R83L) when all three glycosylation sites were substituted. Tunicamycin prevented OCTN2 glycosylation, but it did not impair maturation to the plasma membrane. These results indicate that OCTN2 is physiologically glycosylated and that the P46S and R83L substitutions impair this process. Glycosylation does not affect maturation of OCTN2 transporters to the plasma membrane, but the 3 asparagines that are normally glycosylated are located in a region important for substrate recognition and turnover rate.     

Muscle contraction increases carnitine uptake via translocation of OCTN2

Since carnitine plays an important role in fat oxidation, influx of carnitine could be crucial for muscle metabolism. OCTN2 (SLC22A5), a sodium-dependent solute carrier, is assumed to transport carnitine into skeletal muscle cells. Acute regulation of OCTN2 activity in rat hindlimb muscles was investigated in response to electrically induced contractile activity. The tissue uptake clearance (CL(uptake)) of l-[(3)H]carnitine during muscle contraction was examined in vivo using integration plot analysis. The CL(uptake) of [(14)C]iodoantipyrine (IAP) was also determined as an index of tissue blood flow. To test the hypothesis that increased carnitine uptake involves the translocation of OCTN2, contraction-induced alteration in the subcellular localization of OCTN2 was examined. The CL(uptake) of l-[(3)H]carnitine in the contracting muscles increased 1.4-1.7-fold as compared to that in the contralateral resting muscles (p<0.05). The CL(uptake) of [(14)C]IAP was much higher than that of l-[(3)H]carnitine, but no association between the increase in carnitine uptake and blood flow was obtained. Co-immunostaining of OCTN2 and dystrophin (a muscle plasma membrane marker) showed an increase in OCTN2 signal in the plasma membrane after muscle contraction. Western blotting showed that the level of sarcolemmal OCTN2 was greater in contracting muscles than in resting muscles (p<0.05). The present study showed that muscle contraction facilitated carnitine uptake in skeletal muscles, possibly via the contraction-induced translocation of its specific transporter OCTN2 to the plasma membrane.     

Characterization of organic cation/carnitine transporter family in human sperm

Spermatozoan maturation, motility, and fertility are, in part, dependent upon the progressive increase in epididymal and spermatozoal carnitine, critical for mitochondrial fatty acid oxidation, as sperm pass from the caput to the cauda of the epididymis. We demonstrate that the organic cation/carnitine transporters, OCTN1, OCTN2, and OCTN3, are expressed in sperm as three distinct proteins with an expected molecular mass of 63 kDa, using Western blot analysis and our transporter-specific antibodies. Carnitine uptake studies in normal control human sperm samples further support the presence of high-affinity (OCTN2) carnitine uptake (K(m) of 3.39+/-1.16 microM; V(max) of 0.23+/-0.14 pmol/min/mg sperm protein; and mean+/-SD; n=12), intermediate-affinity (OCTN3) carnitine uptake (K(m) of 25.9+/-14.7 microM; V(max) of 1.49+/-1.03 pmol/min/mg protein; n=26), and low-affinity (OCTN1) carnitine uptake (K(m) of 412.6+/-191 microM; V(max) of 32.7+/-20.5 pmol/min/mg protein; n=18). Identification of individuals with defective sperm carnitine transport may provide potentially treatable etiologies of male infertility, responsive to L-carnitine supplementation.     

A candidate mouse model for Hartnup Disorder deficient in neutral amino acid transport

The mutant mouse strain HPH2 (hyperphenylalaninemia) was isolated after N-ethyl-N-nitrosourea (ENU) mutagenesis on the basis of delayed plasma clearance of an injected load of phenylalanine. Animals homozygous for the recessive hph2 mutation excrete elevated concentrations of many of the neutral amino acids in the urine, while plasma concentrations of these amino acids are normal. In contrast, mutant homozygotes excrete normal levels of glucose and phosphorus. These data suggest an amino acid transport defect in the mutant, confirmed in a small reduction in normalized values of ¹⁴C-labeled glutamine uptake by kidney cortex brush border membrane vesicles (BBMV). The hyperaminoaciduria pattern is very similar to that of Hartnup Disorder, a human amino acid transport defect. A subset of Hartnup Disorder cases also show niacin deficiency symptoms, which are thought to be multifactorially determined. Similarly, the HPH2 mouse exhibits a niacin-reversible syndrome that is modified by diet and by genetic background. Thus, HPH2 provides a candidate mouse model for the study of Hartnup Disorder, an amino acid transport deficiency and a multifactorial disease in the human. Received: 16 May 1996 / Accepted: 25 September 1996