Reticulon Research Articles

Transmembrane protein TMEM170A is a newly discovered regulator of ER and nuclear envelope morphogenesis in human cells.

ABSTRACTThe mechanism of endoplasmic reticulum (ER) morphogenesis is incompletely understood. ER tubules are shaped by the reticulons (RTNs) and DP1/Yop1p family members, but the mechanism of ER sheet formation is much less clear. Here, we characterize TMEM170A, a human transmembrane protein, which localizes in ER and nuclear envelope membranes. Silencing or overexpressing TMEM170A in HeLa K cells alters ER shape and morphology. Ultrastructural analysis reveals that downregulation of TMEM170A specifically induces tubular ER formation, whereas overexpression of TMEM170A induces ER sheet formation, indicating that TMEM170A is a newly discovered ER-sheet-promoting protein. Additionally, downregulation of TMEM170A alters nuclear shape and size, decreases the density of nuclear pore complexes (NPCs) in the nuclear envelope and causes either a reduction in inner nuclear membrane (INM) proteins or their relocalization to the ER. TMEM170A interacts with RTN4, a member of the reticulon family; simultaneous co-silencing of TMEM170A and RTN4 rescues ER, NPC and nuclear-envelope-related phenotypes, implying that the two proteins have antagonistic effects on ER membrane organization, and nuclear envelope and NPC formation.

Dysfunctional tubular endoplasmic reticulum constitutes a pathological feature of Alzheimer's disease.

Pathological features in Alzheimer’s brains include mitochondrial dysfunction and dystrophic neurites (DNs) in areas surrounding amyloid plaques. Using a mouse model that overexpresses reticulon 3 (RTN3) and spontaneously develops age-dependent hippocampal DNs, here we report that DNs contain both RTN3 and REEPs, topologically similar proteins that can shape tubular endoplasmic reticulum (ER). Importantly, ultrastructural examinations of such DNs revealed gradual accumulation of tubular ER in axonal termini, and such abnormal tubular ER inclusion is found in areas surrounding amyloid plaques in biopsy samples from AD brains. Functionally, abnormally clustered tubular ER induces enhanced mitochondrial fission in the early stages of DN formation and eventual mitochondrial degeneration at later stages. Furthermore, such DNs are abrogated when RTN3 is ablated in aging and AD mouse models. Hence, abnormally clustered tubular ER can be pathogenic in brain regions: disrupting mitochondrial integrity, inducing DNs formation and impairing cognitive function in AD and aging brains

A genome-wide association study for colorectal cancer identifies a risk locus in 14q23.1.

Over 50 loci associated with colorectal cancer (CRC) have been uncovered by genome-wide association studies (GWAS). Identifying additional loci has the potential to help elucidate aspects of the underlying biological processes leading to better understanding of the pathogenesis of the disease. We re-evaluated a GWAS by excluding controls that have family history of CRC or personal history of colorectal polyps, as we hypothesized that their inclusion reduces power to detect associations. This is supported empirically and through simulations. Two-phase GWAS analysis was performed in a total of 16,517 cases and 14,487 controls. We identified rs17094983, a SNP associated with risk of CRC [p = 2.5 × 10(-10); odds ratio estimated by re-including all controls (OR) = 0.87, 95% confidence interval (CI) 0.83-0.91; minor allele frequency (MAF) = 13%]. Results were replicated in samples of African descent (1894 cases and 4703 controls; p = 0.01; OR = 0.86, 95% CI 0.77-0.97; MAF = 16 %). Gene expression data in 195 colon adenocarcinomas and 59 normal colon tissues from two different studies revealed that this locus has genotypes that are associated with RTN1 (Reticulon 1) expression (p = 0.001), a protein-coding gene involved in survival and proliferation of cancer cells which is highly expressed in normal colon tissues but has significantly reduced expression in tumor cells (p = 1.3 × 10(-8)).

RTN1 mediates progression of kidney disease by inducing ER stress.

Identification of new biomarkers and drug targets for chronic kidney disease (CKD) is required for the development of more effective therapy. Here we report an association between expression of reticulon 1 (RTN1) and severity of CKD. An isoform-specific increase in the expression of RTN1A is detected in the diseased kidneys from mice and humans, and correlates inversely with renal function in patients with diabetic nephropathy. RTN1 overexpression in renal cells induces ER stress and apoptosis, whereas RTN1 knockdown attenuates tunicamycin-induced and hyperglycaemia-induced ER stress and apoptosis. RTN1A interacts with PERK through its N-terminal and C-terminal domains, and mutation of these domains prevents this effect on ER stress. Knockdown of Rtn1a expression in vivo attenuates ER stress and renal fibrosis in mice with unilateral ureteral obstruction, and also attenuates ER stress, proteinuria, glomerular hypertrophy and mesangial expansion in diabetic mice. Together, these data indicate that RTN1A contributes to progression of kidney disease by inducing ER stress.

Identification of MANF as a protein interacting with RTN1-C.

Reticulons (RTNs) constitute a family of endoplasmic reticulum (ER)-associated proteins with a reticular distribution. Recently, evidence has shown that they exert a cancer-specific proapoptotic function via interaction or modulation of specific proteins. Such evidence is particularly associated with the RTN1-C family members. In order to explore proteins that interact with RTN1-C, the yeast two-hybrid system and regular molecular biological techniques were used to screen the human fetal brain cDNA library. As a result, seven RTN1-C interacting proteins including Homo sapiens mesencephalic astrocyte-derived neurotrophic factor (MANF) were obtained. The interactions between RTN1-C and its interacting proteins were confirmed by β-galactosidase assay and growth test in selective media. Moreover, the MANF/RTN1-C interaction was verified in vitro by glutathione S-transferase pull-down assay and in vivo by immunoprecipitation assay. By immunofluorescence assay, it was found that MANF co-localized with RTN1-C in the ER. Knockdown of RTN1-C reduced the localization of MANF in the ER. These results provide clues to further explore the function of RTN1-C and MANF in neurodegenerative diseases and cancer.

The endogenous caspase-8 inhibitor c-FLIPL regulates ER morphology and crosstalk with mitochondria.

Components of the death receptor-mediated pathways like caspase-8 have been identified in complexes at intracellular membranes to spatially restrict the processing of local targets. In this study, we report that the long isoform of the cellular FLICE-inhibitory protein (c-FLIPL), a well-known inhibitor of the extrinsic cell death initiator caspase-8, localizes at the endoplasmic reticulum (ER) and mitochondria-associated membranes (MAMs). ER morphology was disrupted and ER Ca2+-release as well as ER-mitochondria tethering was decreased in c-FLIP−/− mouse embryonic fibroblasts (MEFs). Mechanistically, c-FLIP ablation resulted in enhanced basal caspase-8 activation and in caspase-mediated processing of the ER-shaping protein reticulon-4 (RTN4) that was corrected by re-introduction of c-FLIPL and caspase inhibition, resulting in the recovery of a normal ER morphology and ER-mitochondria juxtaposition. Thus, the caspase-8 inhibitor c-FLIPL emerges as a component of the MAMs signaling platforms, where caspases appear to regulate ER morphology and ER-mitochondria crosstalk by impinging on ER-shaping proteins like the RTN4.

Impact of RTN3 deficiency on expression of BACE1 and amyloid deposition.

Reticulon 3 (RTN3) has previously been shown to interact with BACE1 and negatively regulate BACE1 activity. To what extent RTN3 deficiency affects BACE1 activity is an intriguing question. In this study, we aimed to address this by generating RTN3-null mice. Mice with complete deficiency of RTN3 grow normally and have no obviously discernible phenotypes. Morphological analyses of RTN3-null mice showed no significant alterations in cellular structure, although RTN3 is recognized as a protein contributing to the shaping of tubular endoplasmic reticulum. Biochemical analysis revealed that RTN3 deficiency increased protein levels of BACE1. This elevation of BACE1 levels correlated with enhanced processing of amyloid precursor protein at the β-secretase site. We also demonstrated that RTN3 deficiency in Alzheimer's mouse models facilitates amyloid deposition, further supporting an in vivo role of RTN3 in the regulation of BACE1 activity. Since it has been shown that RTN3 monomer is reduced in brains of Alzheimer's patients, our results suggest that long-lasting reduction of RTN3 levels has adverse effects on BACE1 activity and may contribute to Alzheimer's pathogenesis.

RTN4 3'-UTR insertion/deletion polymorphism and susceptibility to non-small cell lung cancer in Chinese Han population.

Nogo protein, encoded by gene reticulon-4 (RTN4), includes three major isoforms by different splicing, named Nogo-A Nogo-B and Nogo-C. Nogo proteins play an important role in the apoptosis of cells, especially in tumor cells. RTN4 single nucleotide polymorphisms (SNPs) can influence the efficiency of transcription and translation thus being related with an individual's predisposition to cancer. The CAA insertion/deletion polymorphism (rs34917480) within RTN4 3'-UTR has been reported to be associated with many cancer types. In order to investigate the relationship between this polymorphism and susceptibility to non-small cell lung cancer (NSCLC) in the Chinese population, we conducted the present case-control study including 411 NSCLC patients and 471 unrelated healthy controls. The genotype distributions were significantly different between cases and controls (p=0.014). We found that the del allele could significantly increase NSCLC risk (ins/ins vs ins/del: p=0.007, OR 1.46, 95%CI=1.11-1.93; dominant model: p=0.004, OR 1.47, 95%CI=1.13-1.92 and allele model: p=0.008, OR 1.35, 95%CI=1.08-1.67). This association was stronger in participants over 60 years old, males and smokers. We therefore conclude that the CAA insertion/deletion polymorphism (rs34917480) contributes to non-small cell lung cancer risk in Chinese population. Age, sex and environmental exposure are also related to carcinogenic effects of rs34917480.

Reticulon 3 interacts with NS4B of the hepatitis C virus and negatively regulates viral replication by disrupting NS4B self-interaction.

The non-structural protein 4B (NS4B) of the hepatitis C virus (HCV) is an endoplasmic reticulum (ER) membrane protein comprising two consecutive amphipathic α-helical domains (AH1 and AH2). Its self-oligomerization via the AH2 domain is required for the formation of the membranous web that is necessary for viral replication. Previously, we reported that the host-encoded ER-associated reticulon 3 (RTN3) protein is involved in the formation of the replication-associated membranes of (+)RNA enteroviruses during viral replication. In this study, we demonstrated that the second transmembrane region of RTN3 competed for, and bound to, the AH2 domain of NS4B, thus abolishing NS4B self-interaction and leading to the downregulation of viral replication. This interaction was mediated by two crucial residues, lysine 52 and tyrosine 63, of AH2, and was regulated by the AH1 domain. The silencing of RTN3 in Huh7 and AVA5 cells harbouring an HCV replicon enhanced the replication of HCV, which was counteracted by the overexpression of recombinant RTN3. The synthesis of viral RNA was also increased in siRNA-transfected human primary hepatocytes infected with HCV derived from cell culture. Our results demonstrated that RTN3 acted as a restriction factor to limit the replication of HCV.

Nogo-B Receptor (NgBR): A New Receptor that Modulates Blood Vessel Formation

Nogo-B is an isoform of reticulon-4 (RTN4) that distributes mainly in the endoplasmic reticulum. Nogo-B binds to its receptor, Nogo-B receptor (NgBR), to modulate blood vessel formation. Animal with Nogo-B knockout is phenotypically normal. NgBR is a type I receptor with a single transmembrane domain that binds Nogo-B and probably other angiogenic factors. NgBR knockout in zebra fish leads to abnormal formation of intersomite vessels suggesting NgBR plays a more important role in the Nogo-B/NgBR signaling pathway. It is reported that NgBR plays a role in dolichol biosynthesis, protein N-glycosylation, stabilizes Niemann-Pick type C2 protein, regulates intracellular cholesterol trafficking, controls blood vessel development, and modulates breast cancer progression. However, the research in NgBR remains in its early stage and needs further exploration.

Glutamate provides a key structural contact between reticulon-4 (Nogo-66) and phosphocholine

Human reticulon 4 (RTN-4) has been identified as the neurite outgrowth inhibitor (Nogo). This protein contains a span of 66 amino acids (Nogo-66) flanked by two membrane helices at the C-terminus. We previously determined the NMR structure of Nogo-66 in a native-like environment and defined the regions of Nogo-66 expected to be membrane embedded. We hypothesize that aromatic groups and a negative charge hyperconserved among RTNs (Glu26) drive the remarkably strong association of Nogo-66 with a phosphocholine surface. Glu26 is an isolated charge with no counterion provided by nearby protein groups. We modeled the docking of dodecylphosphocholine (DPC) with Nogo-66 and found that a lipid choline group could form a stable salt bridge with Glu26 and serve as a membrane anchor point. To test the role of the Glu26 anion in binding choline, we mutated this residue to alanine and assessed the structural consequences, the association with lipid and the affinity for the Nogo receptor. In an aqueous environment, Nogo-66 Glu26Ala is more helical than WT and binds the Nogo receptor with higher affinity. Thus, we can conclude that in the absence of a neutralizing positive charge provided by lipid, the glutamate anion is destabilizing to the Nogo-66 fold. Although the Nogo-66 Glu26Ala free energy of transfer from water into lipid is similar to that of WT, NMR data reveal a dramatic loss of tertiary structure for the mutant in DPC micelles. These data show that Glu26 has a key role in defining the structure of Nogo-66 on a phosphocholine surface. This article is part of a special issue entitled: Interfacially Active Peptides and Proteins. Guest Editors: William C. Wimley and Kalina Hristova.

MRNA levels of BACE1 and its interacting proteins, RTN3 and PPIL2, correlate in human post mortem brain tissue

β-Site amyloid precursor protein cleaving enzyme (BACE1) is the rate-limiting enzyme for production of Aβ peptides, proposed to drive the pathological changes found in Alzheimer’s disease (AD). Reticulon 3 (RTN3) is a negative modulator of BACE1 (β-secretase) proteolytic activity, while peptidylprolyl isomerase (cyclophilin)-like 2 (PPIL2) positively regulated BACE1 gene expression in a cell-based assay. This study aimed to analyze RTN3 and PPIL2 mRNA levels in four brain regions from individuals with AD and controls. BACE1 mRNA had been previously quantified in the samples, as had glial fibrillary acidic protein (GFAP) and neuron-specific enolase (NSE), to track changing cell populations in the tissue. mRNA levels in the human post mortem brain tissue were assayed using quantitative real-time polymerase chain reaction (qPCR) and qbasePLUS, employing validated stably expressed reference genes. No differences in RTN3 or PPIL2 mRNA levels were found in individuals with AD, compared to controls. Both RTN3 and PPIL2 mRNA levels correlated significantly with BACE1 mRNA and all three showed similar disease stage-dependent changes with respect to NSE and GFAP. These findings indicated that the in vitro data demonstrating an effect of PPIL2 on BACE1 expression have functional relevance in vivo. Further research into BACE1-interacting proteins could provide a fruitful approach to the modulation of this protease and consequently Aβ production.

The Role of Reticulons in Neurodegenerative Diseases

Reticulons (RTNs) are a group of membrane-associated proteins mainly responsible for shaping the tubular endoplasmic reticulum network, membrane trafficking, inhibition of axonal growth, and apoptosis. These proteins share a common sequence feature, the reticulon homology domain, which consists of paired hydrophobic stretches that are believed to induce membrane curvature by acting as a wedge in bilayer membranes. RTNs are ubiquitously expressed in all tissues, but each RTN member exhibits a unique expression pattern that prefers certain tissues or even cell types. Recently, accumulated evidence has suggested additional and unexpected roles for RTNs, including those on DNA binding, autophagy, and several inflammatory-related functions. These manifold actions of RTNs account for their ever-growing recognition of their involvement in neurodegenerative diseases like Alzheimer’s disease, amyotrophic lateral sclerosis, multiple sclerosis, as well as hereditary spastic paraplegia. This review summarizes the latest discoveries on RTNs in human pathophysiology, and the engagement of these in neurodegeneration, along with the implications of these findings for a better understanding of the molecular events triggered by RTNs and their potential exploitation as next-generation therapeutics.

Increased Expression of Reticulon 3 in Neurons Leads to Reduced Axonal Transport of β Site Amyloid Precursor Protein-cleaving Enzyme 1

BACE1 is the sole enzyme responsible for cleaving amyloid precursor protein at the β-secretase site, and this cleavage initiates the generation of β-amyloid peptide (Aβ). Because amyloid precursor protein is predominantly expressed by neurons and deposition of Aβ aggregates in the human brain is highly correlated with the Aβ released at axonal terminals, we focused our investigation of BACE1 localization on the neuritic region. We show that BACE1 was not only enriched in the late Golgi, trans-Golgi network, and early endosomes but also in both axons and dendrites. BACE1 was colocalized with the presynaptic vesicle marker synaptophysin, indicating the presence of BACE1 in synapses. Because the excessive release of Aβ from synapses is attributable to an increase in amyloid deposition, we further explored whether the presence of BACE1 in synapses was regulated by reticulon 3 (RTN3), a protein identified previously as a negative regulator of BACE1. We found that RTN3 is not only localized in the endoplasmic reticulum but also in neuritic regions where no endoplasmic reticulum-shaping proteins are detected, implicating additional functions of RTN3 in neurons. Coexpression of RTN3 with BACE1 in cultured neurons was sufficient to reduce colocalization of BACE1 with synaptophysin. This reduction correlated with decreased anterograde transport of BACE1 in axons in response to overexpressed RTN3. Our results in this study suggest that altered RTN3 levels can impact the axonal transport of BACE1 and demonstrate that reducing axonal transport of BACE1 in axons is a viable strategy for decreasing BACE1 in axonal terminals and, perhaps, reducing amyloid deposition.

Nogo/RTN4 isoforms and RTN3 expression protect SH-SY5Y cells against multiple death insults

Among the members of the reticulon (RTN) family, Nogo-A/RTN4A, a prominent myelin-associated neurite growth inhibitory protein, and RTN3 are highly expressed in neurons. However, neuronal cell-autonomous functions of Nogo-A, as well as other members of the RTN family, are unclear. We show here that SH-SY5Y neuroblastoma cells stably over-expressing either two of the three major isoforms of Nogo/RTN4 (Nogo-A and Nogo-B) or a major isoform of RTN3 were protected against cell death induced by a battery of apoptosis-inducing agents (including serum deprivation, staurosporine, etoposide, and H2O2) compared to vector-transfected control cells. Nogo-A, -B, and RTN3 are particularly effective in terms of protection against H2O2-induced increase in intracellular reactive oxygen species levels and ensuing apoptotic and autophagic cell death. Expression of these RTNs upregulated basal levels of Bax, activated Bax, and activated caspase 3, but did not exhibit an enhanced ER stress response. The protective effect of RTNs is also not dependent on classical survival-promoting signaling pathways such as Akt and Erk kinase pathways. Neuron-enriched Nogo-A/Rtn4A and RTN3 may, therefore, exert a protective effect on neuronal cells against death stimuli, and elevation of their levels during injury may have a cell-autonomous survival-promoting function.

Preventing formation of reticulon 3 immunoreactive dystrophic neurites improves cognitive function in mice.

Neuritic dystrophy is one of the important pathological features associated with amyloid plaques in Alzheimer's disease (AD) and age-dependent neuronal dysfunctions. We reported previously that reticulon-3 (RTN3) immunoreactive dystrophic neurites (RIDNs) are abundantly present in the hippocampus of AD patients, in AD mouse models, and in aged wild-type mice. Transgenic mice overexpressing the human RTN3 transgene spontaneously develop RIDNs in their hippocampi, and the formation of RIDNs correlates with the appearance of RTN3 aggregation. To further elucidate whether the formation of RIDNs is reversible, we generated transgenic mice expressing wild-type human RTN3 under the control of a tetracycline-responsive promoter. Treatment with doxycycline for 2 months effectively turned off expression of the human RTN3 transgene, confirming the inducible nature of the system. However, the formation of hippocampal RIDNs was dependent on whether the transgene was turned off before or after the formation of RTN3 aggregates. When transgenic human RTN3 expression was turned off at young age, formation of RIDNs was essentially eliminated compared with the vehicle-treated transgenic mice. More importantly, a fear conditioning study demonstrated that contextual associative learning and memory in inducible transgenic mice was improved if the density of RIDNs was lowered. Additional mechanistic study suggested that a reduction in BDNF levels in transgenic mice might contribute to the reduced learning and memory in transgenic mice overexpressing RTN3. Hence, we conclude that age-dependent RIDNs cannot be effectively cleared once they have formed, and we postulate that successful prevention of RIDN formation should be initiated before RTN3 aggregation.

Reduction of β-Amyloid Accumulation by Reticulon 3 in Transgenic Mice

Inhibition of the β-secretase, BACE1, which cleaves amyloid precursor protein (APP) to produce β-amyloid protein (Aβ), is thought to be a feasible therapeutic strategy for Alzheimer's disease. Reticulon (RTN) proteins such as RTN3 have been identified as membrane proteins that interact with BACE1 and inhibit its Aβ-generating activity. In this study, we investigated whether RTN3 can regulate Aβ production in vivo, using transgenic (Tg) mice expressing APP with Swedish and London mutations (APP Tg mice) and those expressing RTN3; the latter mice showed ~1.4-fold higher expression levels of RTN3 protein in the cerebral cortex than non-Tg controls. We analyzed the brains of single APP Tg and double APP/RTN3 Tg mice at the age of approximately 15 months. The levels of secreted APP-β, a direct BACE1 cleavage product of APP, in Tris-soluble fraction were considerably reduced in the hippocampus and cerebral cortex of APP/RTN3 Tg mice relative to those in APP Tg mice. Immunohistochemical analyses demonstrated that Aβ burden and plaques were significantly (by approximately 50%) decreased in both the hippocampus and cerebral cortex of double Tg mice compared to APP Tg mice. Furthermore, the levels of guanidine-soluble Aβ40 and Aβ42 in these brain regions of APP/RTN3 Tg mice were relatively lower than those in APP Tg mice. These findings indicate that even a small increase in RTN3 expression exerts suppressive effects on amyloidogenic processing of APP and Aβ accumulation through modulation of BACE1 activity in vivo, and suggest that induction of RTN3 might be an effective therapeutic strategy against Alzheimer's disease.

C-FLIPL regulates endoplasmic reticulum morphology and mitochondria-associated membranes functions

Physical and functional interactions between endoplasmic reticulum (ER) and mitochondria take place at the mitochondria-associated membranes (MAMs), ER subdomains at the interface between the two organelles. Protein complexes at MAMs regulate lipid synthesis, Ca2+ signaling and apoptosis 1. Furthermore, they influence both ER and mitochondrial morphology, as their ablation is frequently associated to the dramatic remodeling of these organelles. Here we show that c-FLIPL (cellular FLICE-inhibitory protein, long isoform), mainly known as inhibitor of caspase-8, can be retrieved at the ER and MAMs. c-FLIP ablation in mouse embryonic fibroblasts (MEFs) impairs ER morphology and luminal contiguity, by inducing the proliferation of ER cisternae to the detriment of tubular ER. Furthermore, c-FLIPL controls the ER-mitochondria apposition, as the depletion of this protein physically uncouples the two organelles. Moreover, functionally, c-FLIP ablation lowers the cytosolic Ca2+-increase evoked either by agonists stimulation or by passive ER discharge and increases the resistance of c-FLIP-/- cells to ER stress-induced apoptosis. We also report that c-FLIP-/- cells show an increased caspase-mediated cleavage of the ER-shaping protein Reticulon-4 (which is a well-known regulator of ER biogenesis and morphology), at both basal level and upon TNFα-dependent apoptosis. In agreement with these findings, we finally show that c-FLIP absence enhances basal caspase-8 activation in c-FLIP-/- MEFs and that caspase-8 inhibition reverts morphological alterations in ER shape observed in c-FLIP-/- cells, suggesting a novel role for caspase-8 and c-FLIPL as regulators of ER functions and ER-mitochondria crosstalk.

Candidate Human Genetic Polymorphisms and Severe Malaria in a Tanzanian Population

Human genetic background strongly influences susceptibility to malaria infection and progression to severe disease and death. Classical genetic studies identified haemoglobinopathies and erythrocyte-associated polymorphisms, as protective against severe disease. High throughput genotyping by mass spectrometry allows multiple single nucleotide polymorphisms (SNPs) to be examined simultaneously. We compared the prevalence of 65 human SNP's, previously associated with altered risk of malaria, between Tanzanian children with and without severe malaria. Five hundred children, aged 1–10 years, with severe malaria were recruited from those admitted to hospital in Muheza, Tanzania and compared with matched controls. Genotyping was performed by Sequenom MassArray, and conventional PCR was used to detect deletions in the alpha-thalassaemia gene. SNPs in two X-linked genes were associated with altered risk of severe malaria in females but not in males: heterozygosity for one or other of two SNPs in the G6PD gene was associated with protection from all forms of severe disease whilst two SNPs in the gene encoding CD40L were associated with respiratory distress. A SNP in the adenyl cyclase 9 (ADCY9) gene was associated with protection from acidosis whilst a polymorphism in the IL-1α gene (IL1A) was associated with an increased risk of acidosis. SNPs in the genes encoding IL-13 and reticulon-3 (RTN3) were associated with increased risk of cerebral malaria. This study confirms previously known genetic associations with protection from severe malaria (HbS, G6PD). It identifies two X-linked genes associated with altered risk of severe malaria in females, identifies mutations in ADCY9, IL1A and CD40L as being associated with altered risk of severe respiratory distress and acidosis, both of which are characterised by high serum lactate levels, and also identifies novel genetic associations with severe malaria (TRIM5) and cerebral malaria(IL-13 and RTN3). Further studies are required to test the generality of these associations and to understand their functional consequences.

RNT4 3′-UTR Insertion/Deletion Polymorphisms Are Not Associated with Atrial Septal Defect in Chinese Han Population: A Brief Communication

Atrial septal defect (ASD) is a common type of congenital heart disease, which is defined as any communication through atrial septum. Several studies have revealed that genetic factors may influence the susceptibility of ASD. Recent studies have shown that reticulon 4 (RTN4) gene might be involved in some processes relevant to heart development, such as regulation of cell migration and vascular remodeling. This study aimed to evaluate RTN4 gene polymorphisms of CAA and TATC insertion/deletion in relation to the risk of ASD in Chinese Han population. A total of 175 ASD patients and 308 unrelated healthy controls were successfully investigated. The polymorphisms of patients were determined by polymerase chain reaction-polyacrylamide gel electrophoresis. There was no significant difference in the allele frequencies of CAA and TATC insertion/deletion in RNT4 gene between ASD patients and controls. The same results were seen in their genotypes. The present study suggests that CAA and TATC insertion/deletion polymorphisms of RNT4 gene may not be a useful marker to predict the susceptibility of ASD in Chinese Han population.