Mitochondrial DNA Analysis of Mongolian Populations and Implications for the Origin of New World Founders

High levels of mitochondrial DNA (mtDNA) diversity were determined for Mongolian populations, represented by the Mongol-speaking Khalkha and Dariganga. Although 103 samples were collected across Mongolia, low levels of genetic substructuring were detected, reflecting the nomadic lifestyle and relatively recent ethnic differentiation of Mongolian populations. mtDNA control region I sequence and seven additional mtDNA polymorphisms were assayed to allow extensive comparison with previous human population studies. Based on a comparative analysis, we propose that indigenous populations in east Central Asia represent the closest genetic link between Old and New World populations. Utilizing restriction/deletion polymorphisms, Mongolian populations were found to carry all four New World founding haplogroups as defined by WALLACE and coworkers. The ubiquitous presence of the four New World haplogroups in the Americas but narrow distribution across Asia weakens support for GREENBERG and coworkers' theory of New World colonization via three independent migrations. The statistical and geographic scarcity of New World haplogroups in Asia makes it improbable that the same four haplotypes would be drawn from one geographic region three independent times. Instead, it is likely that founder effects manifest throughout Asia and the Americas are responsible for differences in mtDNA haplotype frequencies observed in these regions.     

Single-amino-acid deletion in the RYR1 gene, associated with malignant hyperthermia susceptibility and unusual contraction phenotype

Malignant hyperthermia (MH) is an anesthetic-drug-induced, life-threatening hypermetabolic syndrome caused by abnormal calcium regulation in skeletal muscle. Often inherited as an autosomal dominant trait, MH has linkage to 30 different mutations in the RYR1 gene, which encodes a calcium-release-channel protein found in the sarcoplasmic reticulum membrane in skeletal muscle. All published RYR1 mutations exclusively represent single-nucleotide changes. The present report documents, in exon 44 of RYR1 in two unrelated, MH-susceptible families, a 3-bp deletion that results in deletion of a conserved glutamic acid at position 2347. This is the first deletion, in RYR1, found to be associated with MH susceptibility. MH susceptibility was confirmed among some family members by in vitro diagnostic pharmacological contracture testing of biopsied skeletal muscle. Although a single-amino-acid deletion appears to be a subtle change in the protein, the deletion of Glu2347 from RYR1 produces an unusually large electrically evoked contraction tension in MH-positive individuals, suggesting that this deletion produces an alteration in skeletal-muscle calcium regulation, even in the absence of pharmacological agents.     

Allelic variation at alcohol metabolism genes (<Emphasis Type="Italic">ADH1B</Emphasis>,<Emphasis Type="Italic"> ADH1C</Emphasis>,<Emphasis Type="Italic"> ALDH2</Emphasis>) and alcohol dependence in an American Indian population

Enzymes encoded by two gene families, alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH), mediate alcohol metabolism in humans. Allelic variants have been identified that alter metabolic rates and influence risk for alcoholism. Specifically, ADH1B*47His (previously ADH2-2) and ALDH2-2 have been shown to confer protection against alcoholism, presumably through accumulation of acetaldehyde in the blood and a resultant 'flushing response' to alcohol consumption. In the current study, variants at ADH1B (previously ADH2), ADH1C (previously ADH3), and ALDH2 were assayed in DNA extracts from participants belonging to a Southwest American Indian tribe (n=490) with a high prevalence of alcoholism. Each subject underwent a clinical interview for diagnosis of alcohol dependence, as well as evaluation of intermediate phenotypes such as binge drinking and flushing response to alcohol consumption. Detailed haplotypes were constructed and tested against alcohol dependence and related intermediate phenotypes using both association and linkage analysis. ADH and ALDH variants were also assayed in three Asian and one African population (no clinical data) in order to provide an evolutionary context for the haplotype data. Both linkage and association analysis identified several ADH1C alleles and a neighboring microsatellite marker that affected risk of alcohol dependence and were also related to binge drinking. These data strengthen the support for ADH as a candidate locus for alcohol dependence and suggest further productive study.     

KBTBD13 interacts with Cullin 3 to form a functional ubiquitin ligase

Bone marrow cell (BMC)-derived myofibroblast-like cells have been reported in various organs, including the pancreas. However, the contribution of these cells to pancreatic fibrosis has not been fully discussed. The present study examined the possible involvement of pancreatic stellate cells (PSCs) originating from BMCs in the development of pancreatic fibrosis in a clinically relevant rat model of acute pancreatitis induced by a choline-deficient/ethionine-supplemented (CDE) diet. BMCs from female transgenic mice ubiquitously expressing green fluorescent protein (GFP) were transplanted into lethally irradiated male rats. Once chimerism was established, acute pancreatitis was induced by a CDE diet. Chronological changes in the number of PSCs originating from the donor BMCs were examined using double immunofluorescence for GFP and markers for PSCs, such as desmin and alpha smooth muscle actin (αSMA), 1, 3 and 8 weeks after the initiation of CDE feeding. We also used immunohistochemical staining to evaluate whether the PSCs from the BMCs produce growth factors, such as platelet-derived growth factor (PDGF) and transforming growth factor (TGF) β1. The percentage of BMC-derived activated PSCs increased significantly, peaking after 1 week of CDE treatment (accounting for 23.3±0.9% of the total population of activated PSCs) and then decreasing. These cells produced both PDGF and TGFβ1 during the early stage of pancreatic fibrosis. Our results suggest that PSCs originating from BMCs contribute mainly to the early stage of pancreatic injury, at least in part, by producing growth factors in a rat CDE diet-induced pancreatitis model.     

A protocol comparison for the analysis of heat shock protein A1B +A1538G SNP

Heat shock proteins act as molecular chaperones, assist in peptide maturation, and transport nascent peptides across membranes. One commonly studied single nucleotide polymorphism (SNP) for one of the proteins is HSPA1B (+A1538G). However, several studies of this polymorphism have failed to achieve Hardy–Weinberg equilibrium (HWE) for their sample. We compared various published procedures for analyzing the HSPA1B +A1538G SNP and report reasons for HWE discrepancies. Samples from 141 apparently healthy, physically active, volunteers (99 men and 42 women) were analyzed. The first protocol, initially described by Schröder et al., resulted in a genotypic distribution of 22 GG (15.6%), 119 AG (84.4%), and 0 AA; results were confirmed by reanalysis and sequencing. Two other published protocols, one described by Klausz et al. and another by Fekete et al., were used to confirm these results: both resulted in 22 GG (15.6%), 46 AA (32.6%), and 73 AG (51.7%). Additionally, the results were within HWE and confirmed by sequence analysis. Of the original 119 subjects genotyped as AG by the Schröder protocol, 46 of those were confirmed as AA with the Klausz and Fekete methods. Mixing primers from the Schröder and Klausz protocol resulted in 100% concordance with the data generated by the Klausz and Fekete protocols. Some published data on HSP genotyping deviate from HWE; thus, primers used for analyzing these highly homologous genes must be carefully considered. Our results highlight the importance of reinvestigating data when HWE is not achieved for the HSPA1B, or another, polymorphism.     

High polymorphism at the human melanocortin 1 receptor locus

Variation in human skin/hair pigmentation is due to varied amounts of eumelanin (brown/black melanins) and phaeomelanin (red/yellow melanins) produced by the melanocytes. The melanocortin 1 receptor (MC1R) is a regulator of eu- and phaeomelanin production in the melanocytes, and MC1R mutations causing coat color changes are known in many mammals. We have sequenced the MC1R gene in 121 individuals sampled from world populations with an emphasis on Asian populations. We found variation at five nonsynonymous sites (resulting in the variants Arg67Gln, Asp84Glu, Val92Met, Arg151Cys, and Arg163Gln), but at only one synonymous site (A942G). Interestingly, the human consensus protein sequence is observed in all 25 African individuals studied, but at lower frequencies in the other populations examined, especially in East and Southeast Asians. The Arg163Gln variant is absent in the Africans studied, almost absent in Europeans, and at a low frequency (7%) in Indians, but is at an exceptionally high frequency (70%) in East and Southeast Asians. The MC1R gene in common and pygmy chimpanzees, gorilla, orangutan, and baboon was sequenced to study the evolution of MC1R. The ancestral human MC1R sequence is identical to the human consensus protein sequence, while MC1R varies considerably among higher primates. A comparison of the rates of substitution in genes in the melanocortin receptor family indicates that MC1R has evolved the fastest. In addition, the nucleotide diversity at the MC1R locus is shown to be several times higher than the average nucleotide diversity in human populations, possibly due to diversifying selection.     

Glycyl tRNA synthetase mutations in Charcot-Marie-Tooth disease type 2D and distal spinal muscular atrophy type V

Charcot-Marie-Tooth disease type 2D (CMT2D) and distal spinal muscular atrophy type V (dSMA-V) are axonal peripheral neuropathies inherited in an autosomal dominant fashion. Our previous genetic and physical mapping efforts localized the responsible gene(s) to a well-defined region on human chromosome 7p. Here, we report the identification of four disease-associated missense mutations in the glycyl tRNA synthetase gene in families with CMT2D and dSMA-V. This is the first example of an aminoacyl tRNA synthetase being implicated in a human genetic disease, which makes genes that encode these enzymes relevant candidates for other inherited neuropathies and motor neuron diseases.     

Global patterns of human DNA sequence variation in a 10-kb region on chromosome 1

Human DNA variation is currently a subject of intense research because of its importance for studying human origins, evolution, and demographic history and for association studies of complex diseases. A approximately 10-kb region on chromosome 1, which contains only four small exons (each <155 bp), was sequenced for 61 humans (20 Africans, 20 Asians, and 21 Europeans) and for 1 chimpanzee, 1 gorilla, and 1 orangutan. We found 52 polymorphic sites among the 122 human sequences and 382 variant sites among the human, chimpanzee, gorilla, and orangutan sequences. For the introns sequenced (8,991 bp), the nucleotide diversity (pi) was 0.058% among all sequences, 0.076% among the African sequences, 0.047% among the Asian sequences, and 0.045% among the European sequences. A compilation of data revealed that autosomal regions have, on average, the highest pi value (0.091%), X-linked regions have a somewhat lower pi value (0.079%), and Y-linked regions have a very low pi value (0.008%). The lower polymorphism in the present region may be due to a lower mutation rate and/or selection in the gene containing these introns or in genes linked to this region. The present region and two other 10-kb noncoding regions all show a strong excess of low-frequency variants, indicating a relatively recent population expansion. This region has a low mutation rate, which was estimated to be 0.74 x 10 per nucleotide per year. An average estimate of approximately 12,600 for the long-term effective population size was obtained using various methods; the estimate was not far from the commonly used value of 10,000. Fu and Li's tests rejected the assumption of an equilibrium neutral Wright-Fisher population, largely owing to the high proportion of low-frequency variants. The age of the most recent common ancestor of the sequences in our sample was estimated to be more than 1 Myr. Allowing for some unrealistic assumptions in the model, this estimate would still suggest an age of more than 500,000 years, providing further evidence for a genetic history of humans much more ancient than the emergence of modern humans. The fact that many unique variants exist in Europe and Asia also suggests a fairly long genetic history outside of Africa and argues against a complete replacement of all indigenous populations in Europe and Asia by a small Africa stock. Moreover, the ancient genetic history of humans indicates no severe bottleneck during the evolution of humans in the last half million years; otherwise, much of the ancient genetic history would have been lost during a severe bottleneck. We suggest that both the "Out of Africa" and the multiregional models are too simple to explain the evolution of modern humans.