- Research note
- Open Access
Frequency of Alu insertions within the ACE and PR loci in Northwestern Mexicans
BMC Research Notes volume 10, Article number: 339 (2017)
Presently, non-LTR retrotransposons are the most active mobile elements in the human genome. Among these, Alu elements are highly represented in the modern population. Worldwide, distribution of Alu polymorphisms (insertion/deletion; I/D) shows variability between different populations. Two Alu insertion loci, ACE and PR, are significant biomarkers that have served in several genotype–phenotype association studies. In Mexico, studies concerning the frequency of these biomarkers have been conducted mainly in subpopulations from central and southern regions. Here, we screened a population sample of the northwestern region to gain further knowledge regarding the prevalence of Alu polymorphisms within ACE and PR loci.
For ACE locus, the observed genotype frequencies were 26.5, 51.0 and 22.5% for II, ID, and DD, respectively; and allelic frequencies for I and D were 52 and 48%. Whereas respective genotype frequencies for PR locus were 2.7, 26.5 and 70.8%, and the corresponding allele frequencies were 16 and 84%. Furthermore, the insertion frequency within ACE locus was similar between central, western and northwestern subpopulations, and rather higher in southeastern subpopulation (p < 0.05). Although the occurrence of Alu polymorphisms within PR locus has not been widely examined, the insertion frequency was higher in northwestern subpopulation, as compared with western and southeastern subpopulations (p < 0.05). Based on the frequency of Alu insertions found in ACE and PR loci, subpopulations from the northwestern, western and central regions share a common genetic origin, but apparently not with the subpopulation from the southeastern region, in accordance with the notion that assumes the existence of a broad genomic diversity in the Mexican population. In addition, the high prevalence of Alu insertions reveals their potential application as biomarkers with prognostic value for the associated diseases; e.g., as part of the standard protocols for clinical diagnosis.
Mobile elements can be classified as either DNA transposons or retrotransposons. DNA transposons are currently not mobilizing in the human genome, while retrotransposons have significant mobility. Retrotransposons can be subdivided into LTR and non-LTR elements, which are distinguished by the presence or absence of long terminal repeats. LTR retrotransposons are endogenous retroviruses with very limited activity. By contrast, non-LTR retrotransposons, typified by L1, Alu, and SVA elements, are presently the most active [1,2,3]. Therefore, numerous de novo insertions have resulted in human diseases [3,4,5,6].
Alu elements are one of the most represented in the human genome, being about one million copies (almost 11% of the genome). As active elements in the modern population, new insertions into somatic cells contribute to genomic diversity, gene mutations, and genetic diseases [7, 8]. In addition, the ubiquitous presence of Alu insertions has culminated in their appearance in many genes and transcripts, having a far-reaching influence on gene expression [7, 9, 10]. Moreover, it has been suggested that the most detrimental effect is the interaction between highly homologous elements and their potential to generate deletions, duplications, inversions and other complex genomic rearrangements . Overall, about 0.5% of all human genetic disorders, including some types of cancer, have allegedly resulted from Alu-mediated unequal homologous recombination .
The insertion/deletion (I/D) polymorphism of the angiotensin-converting enzyme (ACE) locus is an important genetic biomarker that has served in numerous genotype–phenotype association studies . ACE plays a key role in the regulation of systemic blood pressure and renal electrolyte homeostasis by converting the inactive angiotensin I into the potent vasoconstrictor and aldosterone-stimulator angiotensin II, and by inactivating the pro-inflammatory vasodilator bradykinin [13, 14]. The I/D polymorphism is distinguished by the presence (insertion) or absence (deletion) of an Alu element within intron 16 of the ACE locus, resulting in three different genotypes: II, ID and DD. Moreover, it seems that the serum concentrations of ACE correlate with the I/D polymorphism (DD > ID > II), suggesting that the levels of circulating enzyme may be determined by the genotype at the ACE locus .
The long-range haplotype, named PROGINS, found in the progesterone receptor (PR) locus comprises an Alu insertion within the intron G (between exon 7 and 8) and two single-nucleotide polymorphisms (SNP): G→T transversion in exon 4, producing a missense mutation (V660L), and C→T transition in exon 5, yielding a silent mutation (H770H). Remarkably, the identification of any of these three alleles uniquely recognizes the presence of the other two . The phenotypic effect of PROGINS is predicted to be due to both the Alu insertion, affecting gene expression and RNA stability, and the V660L substitution, leading to a reduction in the response to progesterone .
Distribution of Alu polymorphisms shows variability among different world populations . In Mexico, studies concerning the frequency of these biomarkers have been conducted mainly in populations of central and southern regions [19,20,21]. Thus, to obtain additional data on the distribution of Alu insertions among other subpopulations, we screened a sample of the northwestern region. Here, we report the prevalence of Alu polymorphisms within ACE and PR loci, two genomic variations which presumably can lead to diseases in humans. Moreover, these biomarkers offer the possibility of being translated to clinical practice after a thorough validation of the genotype–phenotype association.
One hundred and forty-seven samples were available from a DNA biobank collected in a previous study . Any personal data was removed from the sample tube prior to conduct this study; e.g., ensuring individual privacy and autonomy . All samples were assayed for each Alu polymorphism; e.g., ACE or PR. Each segment comprising the insertion was amplified by polymerase chain reactions (PCR) using locus-specific primers (Table 1). Although these assays are sensitive and well documented, precision and reproducibility were ensured by re-typing randomly selected samples throughout the study.
Typical PCR amplifications were performed in a total volume of 0.02 mL of 1X Taq Mix (Qiagen) containing 25 pmol of each primer and 100 ± 40 ng of template DNA. Thermal cycling conditions were as follows: an initial denaturation step (2 min at 94 °C), followed by 35 cycles of exponential amplification (20 s at 94 °C, 20 s at 50 °C, and 40 s at 72 °C), and a final elongation step (7 min at 72 °C). PCR products were separated by gel electrophoresis (2% agarose) and stained with ethidium bromide. The I/D polymorphisms were determined by visual discrimination of each fragment length (Table 1).
The observed values for allele and genotype frequencies were obtained by direct counting, while the expected value for genotype frequency was calculated according to the Hardy–Weinberg (HW) model. HW equilibrium was verified by the goodness-of-fit test χ2 with a significant confidence value of p < 0.05.
Since a significant percentage (around 0.5%) of all human genetic disorders might result from Alu-mediated unequal homologous recombination , the discovery and characterization of the insertion sites are essential in narrowing down the cause of such diseases . Moreover, as Alu elements are the most abundant interspersed repeats in the human genome , the distribution and classification of widespread variants are decisive to identify novel pathogenic insertions [24, 25].
We have reported the prevalence of three pharmacogenetic traits in northwestern Mexicans . To gain additional data regarding other biomarkers, a genetic screening was performed to determine the frequency of Alu polymorphisms within ACE and PR loci (Table 2). For ACE locus, the observed genotype frequencies for II, ID and DD were 26.5, 51.0 and 22.5%; and the frequencies for I and D alleles were 52 and 48%; respectively. Whereas for PR locus, the observed frequencies for II, ID and DD genotypes were 2.7, 26.5 and 70.8%; and the allele frequencies were 16 and 84% for I and D; respectively. Furthermore, distribution of polymorphic genotypes was found to be in Hardy–Weinberg equilibrium (p > 0.05).
Through a comparative analysis (Table 3), we found that the frequency of Alu insertions within ACE locus was quite similar between central, western and northwestern subpopulations, but significantly different in a southeastern subpopulation (p < 0.05), suggesting distinct genetic origins amongst current Mexican subpopulations. Conversely, although the occurrence of Alu polymorphisms within PR locus has not been widely examined, we found that the frequency of insertions was significantly higher in a northwestern subpopulation, as compared with western and southeastern subpopulations (p < 0.05), supporting the previously mentioned notion.
Because of their transposition activity, Alu elements represent a significant source of genomic variation . Their importance becomes highlighted by the potential association with genetic instability, one of the major causes of human diseases such as cancer . In addition, several studies have demonstrated their ability to modulate gene expression at the post-transcriptional level . From the evolutionary standpoint, the Alu elements can be regarded as fixed or polymorphic. Fixed elements are evolutionarily older and present throughout the population, while those that are polymorphic are the result of recent retrotransposition events and can be found in a subset of individuals in the population. Since the prevalence of polymorphic alleles may vary between populations of different origin , genotype screening is essential identifying individuals carrying insertions (e.g., homozygous and heterozygous), especially when the genotype–phenotype association has been well established, because those individuals could be at risk of developing the condition associated [7, 24, 29, 30].
In Mexican populations, the ancestral genetic contribution exhibits regional fluctuations . Although numerous studies have been conducted in different regions, a few have been performed in the northwestern region. Since polymorphic Alu elements are reliable biomarkers [32, 33], we used those within ACE and PR loci to determine the occurrence of I/D polymorphism in the northwestern subpopulation. Our results showed that the frequency of Alu insertions found in the examined subpopulation is quite comparable to the observed in western and central subpopulations, but it is significantly different from that reported in southeastern subpopulation (p < 0.05). This is consistent with the notion that presumes the existence of a broad genomic diversity in the Mexican population [31, 34].
The high prevalence of Alu insertions in current Mexican population reveals their potential application as biomarkers with a prognostic value for the associated diseases; e.g., as part of the standard protocols for laboratory diagnosis. However, this notion should be properly assessed through genotype–phenotype association studies to adequately fulfill clinical purposes.
long terminal repeats
- ACE :
angiotensin converting enzyme
- PR :
single nucleotide polymorphisms
Cordaux R, Batzer MA. The impact of retrotransposons on human genome evolution. Nat Rev Genet. 2009;10(10):691–703.
Muñoz-López M, García-Pérez JL. DNA transposons: nature and applications in genomics. Curr Genom. 2010;11(2):115–28.
Solyom S, Kazazian HH. Mobile elements in the human genome: implications for disease. Genome Med. 2012;4(2):12.
O’Donnell KA, Burns KH. Mobilizing diversity: transposable element insertions in genetic variation and disease. Mob DNA. 2010;1:21.
Ayarpadikannan S, Kim H-S. The impact of transposable elements in genome evolution and genetic instability and their implications in various diseases. Genom Inform. 2014;12(3):98–104.
Hancks DC, Kazazian HH. Roles for retrotransposon insertions in human disease. Mob DNA. 2016;7:9.
Deininger P. Alu elements: know the SINEs. Genome Biol. 2011;12(12):236.
Dridi S. Alu mobile elements: from junk DNA to genomic gems. Scientifica. 2012;2012:545328.
Pandey R, Mukerji M. From ‘JUNK’ to just unexplored noncoding knowledge: the case of transcribed Alus. Brief Funct Genom. 2011;10(5):294–311.
Mandal AK, Pandey R, Jha V, Mukerji M. Transcriptome-wide expansion of non-coding regulatory switches: evidence from co-occurrence of Alu exonization, antisense and editing. Nucleic Acids Res. 2013;41(4):2121–37.
Cook GW, Konkel MK, Walker JA, et al. A comparison of 100 human genes using an Alu element-based instability model. PLoS ONE. 2013;8(6):e65188.
Morales ME, White TB, Streva VA, Defreece CB, Hedges DJ, Deininger PL. The contribution of Alu elements to mutagenic DNA double-strand break repair. PLoS Genet. 2015;11(3):e1005016.
Wang K, Li Y, Dai C, et al. Characterization of the relationship between APOBEC3B deletion and ACE Alu insertion. PLoS ONE. 2013;8(5):e64809.
Demurtas P, Orrù G, Coni P, et al. Association between the ACE insertion/deletion polymorphism and pterygium in Sardinian patients: a population based case–control study. BMJ Open. 2014;4(10):e005627.
Rigat B, Hubert C, Alhenc-gelas F, Cambien F, Corvol P, Soubrier F. An insertion/deletion polymorphism in the angiotensin I-converting enzyme gene accounting for half the variance of serum enzyme levels. J Clin Investig. 1990;86(4):1343–6.
Pearce CL, Hirschhorn JN, Wu AH, et al. Clarifying the PROGINS allele association in ovarian and breast cancer risk: a haplotype-based analysis. J Natl Cancer Inst. 2005;97(1):51–9.
Romano A, Delvoux B, Fischer DC, Groothuis P. The PROGINS polymorphism of the human progesterone receptor diminishes the response to progesterone. J Mol Endocrinol. 2007;38(1–2):331–50.
Watkins WS, Rogers AR, Ostler CT, et al. Genetic variation among world populations: inferences from 100 Alu insertion polymorphisms. Genome Res. 2003;13(7):1607–18.
Vargas-Alarcón G, Hernández-Pacheco G, Rodríguez-Pérez JM, et al. Angiotensin-converting enzyme gene (ACE) insertion/deletion polymorphism in Mexican populations. Hum Biol. 2003;75(6):889–96.
Herrera RJ, Rojas DP, Terreros MC. Polymorphic Alu insertions among Mayan populations. J Hum Genet. 2007;52(2):129–42.
Gallegos-Arreola MP, Figuera LE, Flores-Ramos LG, Puebla-Pérez AM, Zúñiga-González GM. Association of the Alu insertion polymorphism in the progesterone receptor gene with breast cancer in a Mexican population. Arch Med Sci. 2015;11(3):551–60.
Ramos MA, Mares RE, Avalos ED, et al. Pharmacogenetic screening of N-acetyltransferase 2, thiopurine s-methyltransferase, and 5,10-methylene-tetrahydrofolate reductase polymorphisms in Northwestern Mexicans. Genet Test Mol Biomark. 2011;15(5):351–5.
Bathe OF, McGuire AL. The ethical use of existing samples for genome research. Genet Med. 2009;11(10):712–5.
Hormozdiari F, Alkan C, Ventura M, et al. Alu repeat discovery and characterization within human genomes. Genome Res. 2011;21(6):840–9.
Gonzaga-Jauregui C, Lupski JR, Gibbs RA. Human genome sequencing in health and disease. Annu Rev Med. 2012;63:35–61.
Zhang W, Edwards A, Fan W, Deininger P, Zhang K. Alu distribution and mutation types of cancer genes. BMC Genom. 2011;12:157.
Häsler J, Strub K. Alu elements as regulators of gene expression. Nucleic Acids Res. 2006;34(19):5491–7.
Hedges DJ, Belancio VP. Restless genomes humans as a model organism for understanding host-retrotransposable element dynamics. Adv Genet. 2011;73:219–62.
Cardelli M, Marchegiani F, Provinciali M. Alu insertion profiling: array-based methods to detect Alu insertions in the human genome. Genomics. 2012;99(6):340–6.
Qian Y, Kehr B, Halldórsson BV. PopAlu: population-scale detection of Alu polymorphisms. PeerJ. 2015;3:e1269.
Martinez-Fierro ML, Beuten J, Leach RJ, et al. Ancestry informative markers and admixture proportions in Northeastern Mexico. J Hum Genet. 2009;54(9):504–9.
Batzer MA, Deininger PL. Alu repeats and human genomic diversity. Nat Rev Genet. 2002;3(5):370–9.
Athanasiadis G, Esteban E, Via M, et al. The X chromosome Alu insertions as a tool for human population genetics: data from European and African human groups. Eur J Hum Genet. 2007;15(5):578–83.
Silva-Zolezzi I, Hidalgo-Miranda A, Estrada-Gil J, et al. Analysis of genomic diversity in Mexican Mestizo populations to develop genomic medicine in Mexico. Proc Natl Acad Sci USA. 2009;106(21):8611–6.
Topete-Reyes JF, Soto-Vargas J, Morán-Moguel MC, et al. Insertion/deletion polymorphism of the angiotensin-converting enzyme gene in lupus nephritis among Mexicans. Immunopharmacol Immunotoxicol. 2013;35(1):174–80.
Valdez-Velazquez LL, Quintero-Ramos A, Perez SA, et al. Genetic polymorphisms of the renin–angiotensin system in preterm delivery and premature rupture of membranes. J Renin Angiotensin Aldosterone Syst. 2007;8(4):160–8.
Alvarez-Aguilar C, Enríquez-Ramírez ML, Figueroa-Nuñez B, et al. Association between angiotensin-1 converting enzyme gene polymorphism and the metabolic syndrome in a Mexican population. Exp Mol Med. 2007;39(3):327–34.
Avila-Vanzzini N, Posadas-Romero C, del Gonzalez-Salazar M, et al. The ACE I/D polymorphism is associated with nitric oxide metabolite and blood pressure levels in healthy Mexican men. Arch Cardiol Mex. 2015;85(2):105–10.
Rupert JL, Kidd KK, Norman LE, Monsalve MV, Hochachka PW, Devine DV. Genetic polymorphisms in the renin–angiotensin system in high-altitude and low-altitude Native American populations. Ann Hum Genet. 2003;67(Pt 1):17–25.
REM and MAR conceived and designed the study. HPN and LHS performed the experiments and analyzed the data. MAR wrote the first draft of the manuscript. HPN, LHS, and REM proofread the manuscript. All authors read and approved the final manuscript.
REM and MAR are national researchers (SNI-CONACYT) and members of the biological-pharmaceutical academic group (Health Sciences, UABC).
The authors declare that they have no competing interests.
Availability of data and materials
All data generated or analyzed during this study are included in this published article.
Consent for publication
Ethics approval and consent to participate
The study received ethical clearance from the Research Board of the Department of Research and Graduate Studies, Autonomous University of Baja California, Tijuana (Approval No. 300/1552). No experiments were conducted on animals or human subjects. The human material used (i.e., a DNA biobank) was collected in a previous study . Further approval from a bioethics committee was not required. All included participants voluntarily signed a written informed consent, which also claimed protection for individual privacy by removing any personal data from the sample tube before executing any analysis (current or subsequent). Medical research was performed in accordance with the Declaration of Helsinki, as explicitly declared in the ethical code of the approved protocol.
The study was supported in part by grants from the Mexican Council for Science and Technology, CONACyT (CB-2010/01-155714 and SSA/IMSS/ISSSTE-2011/01-161544).
About this article
Cite this article
Navarrete, H.P., Soler, L.H., Mares, R.E. et al. Frequency of Alu insertions within the ACE and PR loci in Northwestern Mexicans. BMC Res Notes 10, 339 (2017). https://doi.org/10.1186/s13104-017-2673-y
- Human polymorphisms
- Alu insertions
- Mexican population
- Genomic diversity