Skip Navigation


Rheumatology Advance Access originally published online on July 11, 2006
Rheumatology 2007 46(2):220-226; doi:10.1093/rheumatology/kel210
This Article
Right arrow Abstract Freely available
Right arrow FREE Full Text (PDF) Freely available
Right arrowOA All Versions of this Article:
46/2/220    most recent
kel210v1
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in ISI Web of Science
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Add to My Personal Archive
Right arrow Download to citation manager
Right arrow Search for citing articles in:
ISI Web of Science (3)
Right arrow Disclaimer
Google Scholar
Right arrow Articles by Kong, E. K.-P.
Right arrow Articles by Lau, Y.-L.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Kong, E. K.-P.
Right arrow Articles by Lau, Y.-L.
Related Collections
Right arrow Systemic Lupus Erythematosus and Autoimmunity
Right arrow Immunogenetics
Social Bookmarking
Add to CiteULike   Add to Connotea   Add to Del.icio.us  
What's this?

© The Author 2006. Published by Oxford University Press on behalf of the British Society for Rheumatology. All rights reserved. For Permissions, please email: journals.permissions@oxfordjournals.org
The online version of this article has been published under an open access model. Users are entitled to use, reproduce, disseminate, or display the open access version of this article for non-commercial purposes provided that: the original authorship is properly and fully attributed; the Journal and Oxford University Press are attributed as the original place of publication with the correct citation details given; if an article is subsequently reproduced or disseminated not in its entirety but only in part or as a derivative work this must be clearly indicated. For commercial re-use, please contact journals.permissions@oxfordjournals.org

p21 gene polymorphisms in systemic lupus erythematosus

E. K.-P. Kong, W.-P. Chong, W. H.-S. Wong, C.-S. Lau1, T.-M. Chan1, P. K.-M. Ng2, Y.-Q. Song2, W. Mak2 and Y.-L. Lau

Department of Paediatrics and Adolescent Medicine, 1Department of Medicine and 2Department of Genome Research Centre, The Hong Kong Jockey Club Research Center, The University of Hong Kong, Pokfulam, Hong Kong SAR, China

Correspondence to: Yu-Lung Lau, MD, Department of Paediatrics and Adolescent Medicine, Queen Mary Hospital, The University of Hong Kong, Pokfulam, Hong Kong SAR, China. E-mail: lauylung{at}hkucc.hku.hk


   Abstract
Top
 Abstract
Introduction
 Materials and methods
 Results
 Discussion
 Acknowledgements
 References
 
Objective. Cyclin-dependent kinase inhibitor 1A (p21) is a negative regulator in the cell cycle. Development of sex-linked lupus-like syndrome in p21–/– mice and reduced p21 gene expression in patients with systemic lupus erythematosus (SLE) compared with those in healthy controls suggested that p21 is a susceptibility gene of SLE. We investigated the same by a case-control association study.

Methods. Six single nucleotide polymorphisms, p21US G/A, p21DS C/A, p21-1022 G/A, p21C31 C/A, p21In2 G/C and p21UTR T/C, were genotyped in 516 SLE patients and 693 healthy controls. Association of genotypes and alleles with disease, disease phenotypes, haplotypes construction, linkage disequilibrium analysis and p21 mRNA expression were performed.

Results. We found a significant association of p21US A allele (OR = 0.23, 95% CI: 0.14–0.38, P < 0.001) and p21-1022 A allele (OR = 1.95, 95% CI: 1.37–2.78, P < 0.001) with SLE. We identified significant differences in the frequencies of haplotypes ht1-ACACCC, which contains p21US A allele, and ht2-GCACCC, which contains p21-1022 A allele, between SLE patients and controls (P < 0.0001). Besides, the p21US GA was associated with SLE patients suffering from arthritis (P = 0.003). We also observed differential p21 mRNA expressions among different genotypes of p21US and p21-1022 which were statistically significant.

Conclusion. Our results suggested that the p21US A allele and p21-1022 A allele were both associated with the development of SLE, and the p21US A allele was associated with arthritis in SLE patients.

KEY WORDS: SLE, p21, Susceptibility, Single nucleotide polymorphism, Haplotype


   Introduction
Top
 Abstract
 Introduction
Materials and methods
 Results
 Discussion
 Acknowledgements
 References
 
Cyclin-dependent kinase inhibitor 1A (also known as p21, WAF1 or CDKN1A) is a negative regulator of cyclin dependent kinases (CDKs) [1]. It is a 21 kDa cell cycle regulatory protein that forms quaternary complexes with the entire cyclin/CDK holoenzyme and thereby inhibiting the progression from the G1 to the S phase of the cell cycle [2–5]. It can also bind to the replication factor proliferating cell nuclear antigen (PCNA) and inhibit DNA replication [6]. The p21 gene contains the p53 binding site that is localized at 2.4 kb upstream from the translational start site and expression of p21 gene was found to be inducible by wild-type p53 gene expression [7]. Moreover, several growth factors and cytokines, including interferon (IFN)-{alpha} and -{gamma}, can modify p21 expression [8, 9].

p21–/– mice are deficient in G1 cell cycle arrest, although they develop normally [10]. In addition, cells lacking p21 gene are defective in DNA repair, which might induce impaired regulation in cell proliferation [11]. Inactivating mutations of p21 lead to overexpression and hyperactivation of low-avidity, autoreactive T-cells that are found in abundance in the peripheral lymphoid organs of normal individuals [12]. Indeed, the p21–/– female mice with 129/Sv x C57BL/6 mixed background show development of severe lupus-like diseases followed by early mortality, high levels of anti-dsDNA antibodies, kidney immune complex deposits and glomerulonephritis [13]. However, the p21–/– lupus-prone BXSB mice show inhibition for the development of systemic autoimmunity via the enhancement of the Fas/FasL-mediated activation-induced cell death (AICD) of T- and B-cells [14]. Although these studies show conflicting results, they somehow provide the evidence that p21 is involved in the pathogenesis of systemic lupus erythematosus (SLE), and this discrepancy is probably due to the different genetic background of the mice [13, 14].

The human p21 gene maps on chromosome 6p21.2 [7], which is a susceptibility region for SLE [15, 16]. In addition, reduced p21 expression is observed in peripheral blood lymphocytes and synovial fibroblasts in patients with SLE and rheumatoid arthritis (RA), respectively [17, 18]. In addition, the T-cells of p21–/– mice show enhanced proliferation when compared with T-cells of p21+/+ mice [13, 14, 19]. These indicated p21 deficiency or reduced expression that might contribute to the dysregulated lymphocyte proliferation.

These findings suggested that p21 is a strong candidate gene for the susceptability autoimmune diseases susceptibility, especially for SLE. We therefore aimed to examine whether p21 is associated with SLE in our population. We studied the correlation of six single nucleotide polymorphisms (SNPs) of p21 (GenBank accession number AF497972 [GenBank] ) with SLE in a case-control association study of 1209 Hong Kong Chinese. Moreover, we investigated the correlation between p21 polymorphisms and its mRNA expression.


   Materials and methods
Top
 Abstract
 Introduction
 Materials and methods
Results
 Discussion
 Acknowledgements
 References
 
Study subjects
A total of 516 Hong Kong Chinese patients with SLE recruited from the Queen Mary Hospital were studied. Their mean age was 39 ± 12 yrs (91% female), and they satisfied the revised American College of Rheumatology (formerly, the American Rheumatism Association) criteria for systemic lupus [20, 21]. The study was approved by the Institutional Review Board of the University of Hong Kong/Hospital Authority Hong Kong West Cluster, and all patients gave informed consent. Clinical and serological data, and autoantibody profiles were carefully recorded at the time of diagnosis with SLE. Renal disease was defined as proteinuria of ≥0.5 gm/day or biopsy-proven lupus nephritis. Neurological disorders included psychosis, seizure, organic brain syndrome, aseptic meningitis, depression and cognitive dysfunction. Haematological disorders included haemolytic anaemia, leucopenia, lymphopenia or thrombocytopenia. A total of 693 healthy and ethnically matched blood donors with mean age 30 ± 9 yrs (40% female) from Red Cross served as controls.

Genomic DNA from patients and controls was extracted from ethylenediaminetetraacetic acid (EDTA)-treated whole blood using the DNA Blood Mini Kit (Qiagen, Hilden, Germany) according to the manufacturer's instructions. The DNA samples were then stored at 4°C until used.

Genotyping
SNPs p21In2 and p21UTR were analysed by the polymerase chain reaction (PCR)-restriction fragment length polymorphism (RFLP). A 448 bp fragment of p21In2 and a 480 bp fragment of p21UTR were separately amplified by PCR in a 10 µl reaction mixture as described [22]. The PCR products of p21In2 and p21UTR were then digested with 2 U of ApaL I and PstI, respectively (New England Biolabs, Inc. Beverly, MA, USA), at 37°C overnight, and the fragments were resolved on a 2% agarose gel. For p21In2, the G allele lacks a ApaL I site that is present in the C allele. Therefore, digestion results in fragments of 448 bp for the G allele or 289 and 159 bp for the C allele. For p21UTR, the T allele lacks an Pst I site that is present in the C allele. Therefore, the digestion results in fragments of 480 bp for the T allele or 291 and 189 bp for the C allele. The sequences of primers are shown in Table 1.


View this table:
[in this window]
[in a new window]

  		TABLE 1. Primers used for p21 SNPs genotyping

 
SNPs p21US, p21DS, p21-1022 and p21C31 were genotyped by the MassARRAY system (Sequenom, San Diego, CA). In brief, the samples were amplified in a 6 µl reaction mixture, containing 5 ng genomic DNA, 0.3 pmol each of specific forward and reverse primers, 200 µM of each dNTP, 3.25 mM MgCl2 and 0.2 units of HotStarTaq polymerase (Qiagen, Valencia, CA). The PCR condition included initial hot-start for 15 min at 95°C, 45 cycles of amplification (20 s at 95°C, 30 s at 56°C and 1 min at 72°C) and final extension for 3 min at 72°C. The PCR products were treated with alkaline phosphatase to dephosphorylate residual amplification nucleotides. A mixture of 0.2 µl hME buffer, 0.3 µl shrimp alkaline phosphatase (1 unit/µl, Sequenom) and 1.5 µl ddH2O was added to the PCR products. The reaction solutions were incubated for 20 min at 37°C, followed by 5 min at 85°C to inactivate the enzyme. Mass-extend reactions to determine genotypes were performed in four groups of different terminations according to the design rationale (ddACG, ddACT, ddAGT and ddCGT, respectively). The reaction volume was 10 µl including 1 unit of Thermosequenase (Sequenom), 50 µM of the respective termination mix and 0.6 pmol of each assay specific extension primer. All assays were run with the same thermal cycle conditions: initial denaturation for 2 min at 94°C followed by 55 cycles of extension (5 s at 94°C, 5 s at 52°C and 5 s at 72°C). Products of the mass-extend reactions were desalted and transferred onto a SpectroCHIP by a nanolitre dispenser according to the manufacturer's instructions (Sequenom).

Genotype determination was performed on a MALDI-TOF mass spectrometer (Sequenom). Mass spectrometric data was automatically imported into the SpectroTYPER (Sequenom) database for data analysis including noise normalization and peak area analysis. The expected molecular weights of all relevant peaks were calculated by the MassARRAY AssayDesign Software (Sequenom) before the analysis and were identified from the mass spectrum. In every 96-well plate for assay, there is one well for blank control and five wells for duplicate check on five samples for internal quality control. The sequences of primers are shown in Table 1.

Detection of mRNA expression
Peripheral blood mononuclear cells (PBMCs) (106 cells/ml) from 100 healthy female Red Cross blood donors were cultured with or without 10 µg/ml PHA (Sigma, USA) at 37°C, 5% CO2 for 2 h. The cultured cells were harvested and total RNA was extracted from the cell pellets using the RNeasy mini kit (Qiagen, Hilden, Germany) according to the manufacturer's instructions. cDNA was generated from the extracted total RNA using the TaqMan reverse transcription reagents (Applied Biosystems, USA) according to the manufacturer's instructions. Relative mRNA expression of p21 was performed by a relative quantification reverse transcription (RT)-PCR. A volume of 20 µl PCR mixture containing 20 ng of cDNA, 1 µl 20 x p21 assay mix (Assay-on-demand [Hs00355782_m1], Applied Biosystems, USA), 1 µl 20 x GAPDH mix (Applied Biosystems, USA) and 10 µl TaqMan Universal Master Mix (Applied Biosystems, USA) was used. The real-time PCR was performed in ABI7700 machine (Applied Biosystems) as follows: 5 min at 50°C for decontamination via uracil-N-glycosylase (UNG), 10 min at 95°C for AmpliTaq Gold pre-activation, and was amplified for 50 cycles (15 s at 95°C and 1 min at 60°C). Each sample was run in triplicate.

Statistical analysis
Linkage disequilibrium (LD) and haplotype analysis
LD analysis for p21 SNPs were performed by using chi-square test. The genotype frequencies of all SNPs were tested for Hardy–Weinberg equilibrium (HWE) separately in SLE patients and controls by chi-square test. The multiple-locus haplotypes (hts) comprising of the six SNPs of p21 gene were estimated by using the expectation-maximization (EM) algorithm with 1000 permutations which estimate the maximum-likelihood of the haplotype frequencies for the patients and control separately, then comparing the difference of the haplotype frequencies for the overall subjects.

Association and disease phenotypes analyses
To determine the association of p21 SNPs with SLE patients, a two-step analysis was performed. The genotype frequencies of all six p21 SNPs were compared between SLE patients and controls by a 3 x 2 chi-square test, then multiple logistic regression was used for calculating odds ratios (OR) [95% confidence interval (CI)] and corresponding P-values of different genotype frequencies among SLE patients and controls, adjusting for the sex and age. The association between the clinical features and autoantibody profiles of 393 SLE patients with various genotypes were analysed by using chi-square test. Since all clinical features were recorded at the time of diagnosis, disease duration was not included as a covariable for statistical analysis in clinical features, autoantibody profiles and p21 polymorphisms analyses. All statistical analyses were performed by using SAS, version 8.02 and SAS/Genetics (SAS Institute Inc., NC, USA). Bonferroni correction was used to control the multiple testing effect.

Relative p21 mRNA expression
The analysis of relative quantification was done according to the manufacturer's instructions. The mean fold change in p21 mRNA expression that normalized with GAPDH mRNA expression was calculated by the 2–{Delta}{Delta}Ct method [23]. Ct, threshold cycle, represents the fractional cycle number at which the amount of amplified target reaches a fixed threshold and {Delta}Ct is equal to the difference in Ct for p21 and GAPDH. The {Delta}{Delta}Ct value equals to (Ct,p21 – Ct,GAPDH)S – (Ct,p21 – Ct,GAPDH)C, where S and C indicate the samples with PHA stimulation and without PHA stimulation, respectively. Data were expressed as mean using the scatter dot plots. Kruskal–Wallis test and Mann–Whitney test (GraphPad Prism version 4.0; GraphPad Inc.) were used to detect any significance among the genotypes.


   Results
Top
 Abstract
 Introduction
 Materials and methods
 Results
Discussion
 Acknowledgements
 References
 
HWE and LD
All frequencies of the six polymorphic loci of p21 in SLE patients and controls were in HWE P > 0.05), except the frequency of p21In2 in controls (P < 0.012) and p21DS in SLE patients (P < 0.011). Strong LD was found among the SNPs, but no absolute LD was observed (Table 2). p21 lies in a haplotype block which can be characterized by five haplotype-tagged SNPs according to the HapMap project in Chinese (www.hapmap.org). The two p21 haplotype-tagged SNPs p21c31 (rs1801270) and p21In2 (rs3176352) were included in this study and the other three haplotype-tagged SNPs (rs876581, rs12528248 and rs7761648) lie outside p21.


View this table:
[in this window]
[in a new window]

  		TABLE 2. LD of p21 SNPs in this sudy

 
Association between p21 SNPs and SLE
As illustrated in Table 3, significant differences in allele frequencies of p21US A allele (OR = 0.23; P < 0.001) and p21-1022 A allele (OR = 1.95; P < 0.001) were observed between SLE patients and controls. The remaining SNPs did not show any significant difference between patients and controls. In addition, significant differences in genotype frequencies of p21US, p21-1022, p21In2 and p21UTR were observed among SLE patients and controls (Table 4). Frequency of p21US GA genotype was reduced in SLE patients (OR = 0.23; P < 0.001). Similarly, reduction in genotype frequencies of p21-1022 GG (OR = 0.52; P = 0.001), p21In2 GG (OR = 0.49; P < 0.001) and p21UTR TT (OR = 0.63; P = 0.003) were also observed in SLE patients.


View this table:
[in this window]
[in a new window]

  		TABLE 3. Allele frequencies of p21 SNPs in SLE patients and controls

 

View this table:
[in this window]
[in a new window]

  		TABLE 4. Genotype frequencies of p21 SNPs in SLE patients and controls

 
Haplotype analysis
In theory, a total of 64 (26) possible haplotypes could be constructed from six SNPs. To reduce the complexity, the haplotypes with an overall estimated frequency (haplotype frequency of SLE patients + controls) <3% were not shown (Table 5). Three haplotypes, ht1 (ACACCC), ht2 (GCACCC) and ht3 (GAGAGT), showed significant differences between SLE patients and controls (P < 0.0001), and especially for the ht2, the frequency increased from 0.28% in the controls to 8% in SLE patients (Table 5).


View this table:
[in this window]
[in a new window]

  		TABLE 5. Haplotype analysis of p21 SNPs in SLE patients and controls*

 
Association between genotype and clinical features
All six polymorphisms were analysed for association with clinical features and autoantibody profiles. Only p21US was found to show a significant association, while the remaining SNPs were not (unpublished data). As illustrated in Table 6, the genotype frequency of p21US GA was significantly increased in SLE patients with arthritis (OR = 5.14; P = 0.003) and serositis (OR = 3.86; P = 0.024). After correction by Bonferroni method, the significant P-value should be <0.0056, hence only arthritis remained to have significance (P = 0.003).


View this table:
[in this window]
[in a new window]

  		TABLE 6. Genotype frequencies of p21US of SLE patients with various clinical features and autoantibody profiles

 
Differential p21 mRNA expression for different genotypes of p21US and p21-1022
For p21US, the highest expression was observed for AA genotype (mean = 0.513), followed by AG (mean = 0.491) and GG genotypes (mean = 0.332) for p21US (Fig. 1A). The difference was significant between p21US AG and p21US GG (P = 0.0039) by Mann–Whitney test. For p21-1022, a comparison between AG (mean = 0.288) and GG genotypes (mean = 0.380) alone was available, since AA genotype was not found in the tested samples. These two genotypes also showed differential expression level of p21 mRNA, which was statistically significant (P = 0.0101) (Fig. 1B).


Figure 1
View larger version (27K):
[in this window]
[in a new window]
[Download PowerPoint slide]
  		FIG. 1. Relative quantification of p21 mRNA expression. A–F represent the mean fold change in p21 mRNA expression with different genotypes of p21US, p21DS, p21-1022, p21C31, p21In2 and p21UTR, respectively. The p21 mRNA expression was normalized with that of GAPDH. Only expressions of two genotypes are available for p21-1022 as AA genotype is extremely rare (<1% in controls). P-values >0.05 are not shown. Mean fold change of gene expression is determined by using the 2–{Delta}{Delta}Ct method [23]. n = number of samples.

 

   Discussion
Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
Acknowledgements
 References
 
In the present study, we identified the significant association of p21US A allele with decreased susceptibility to SLE (OR = 0.23, P < 0.001). The haplotype ht1-ACACCC, which contains p21US A allele, was also found to be under-represented in SLE patients (1.89%, 95% CI: 1.06–2.73) when compared with controls (4.48%, 95% CI: 3.39–5.57). Furthermore, we observed the association of p21-1022 A allele with SLE susceptibility (OR = 1.95, P < 0.001). Similarly, the haplotype ht2-GCACCC, which contains p21-1022 A allele, was found to be over-represented in SLE patients (8.00%, 95% CI: 6.34–9.66) when compared with controls (0.28%, 95% CI: 0–0.56). Interestingly, ht1-ACACCC contains both of the protective allele, p21US A, and the risk allele, p21-1022 A, but only the protective effect was observed as the ht1 frequency was significantly reduced in SLE patients (P < 0.001). This implicated that p21US A allele has a dominant effect over the p21-1022 A allele at the haplotype level.

The exact underlying mechanisms that account for the association of p21 SNP in SLE patients are yet to be identified. Both p21US and p21-1022 are localized in the promoter region. One possible mechanism is that these two SNPs may alter the level of p21 expression by modifying the promoter activity. p21US is situated 5 bp upstream from the p53 promoter binding site, the p21US A allele might increase the binding affinity of p53, thus enhance p21 expression [7]. As shown in our p21 mRNA expression assay, individuals with AA or GA genotypes have higher p21 levels than those with GG genotype (Fig. 1A), and the low frequencies of p21US AA and p21US GA in SLE patients may implicate that a high p21 level is a protective factor in the development of SLE. However, p21 expression can be activated by p53-independent pathway [24]. Whether the p53-dependent or p53-independent activation of p21 expression plays an important role in autoimmunity is still elusive, the presence of anti-DNA and anti-p53 antibodies in SLE patients that could functionally block p53 activation [25] suggested that both pathways are probably involved in autoimmunity. Further study is necessary to address this issue. Another promoter SNP, p21-1022, is situated in a potential binding site of E2F (predicted from the TFSEARCH database), which is a strong transcription enhancer of p21 [24, 26]. The substitution of G allele by A allele abolishes the E2F binding site and we demonstrated the risk p21-1022 A allele was correlated to a lower p21 mRNA expression than p21-1022 G allele (Fig. 1C), which further supports the role of p21 in SLE development.

The p21US A allele was associated with increased risk of arthritis in SLE patients, although it was associated with protection against SLE development. These observations were in agreement with the findings of opposite outcomes in the p21–/– mice with different genetic backgrounds as discussed in Introduction [13, 14].

Case-control association study is a powerful tool to identify candidate genes in disease susceptibility [27]. However, statistical power and multiple testing effect should be addressed [28–30]. In this study, a relatively large sample size with over 1000 ethnically matched subjects was investigated so as to achieve adequate statistical power. We also employed Bonferroni method to control the multiple testing effect to avoid type I error. As discussed by Huizinga et al. [30], a further replica study is required with larger independent patient and control groups or family-based method, especially for the SNPs with low allele frequency.

Population stratification is another important issue in case-control association study. In this study, we have recruited homogeneous ethnically matched Hong Kong Chinese subjects. The genotype distribution of the six p21 SNPs in control groups were in HWE (P > 0.05) except the p21In2 with marginal deviation (P < 0.012). To confirm that there is no genotyping error causing this deviation, direct DNA sequencing was performed in 30–50 samples for each SNP and no ambiguous results were obtained. Therefore, this marginal deviation from HWE for p21In2 is much likely due to the multiple testing effect.

In conclusion, we have provided evidence that the p21 gene polymorphisms were associated with SLE, especially SNP p21US and p21-1022. The p21US A allele was identified to be a protective factor in the development of SLE (OR = 0.23) and this allele also showed a dominant effect over the risk associated allele, p21-1022 A allele, at haplotype level. The protective p21US A allele and the risk p21-1022 A allele were associated with high and low p21 mRNA expression respectively, implicating that a high p21 level may reduce the chance of developing SLE. These findings should provide a new insight for the role of cell cycle regulatory proteins in the development of autoimmune diseases.

Formula


   Acknowledgements
Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Acknowledgements
References
 
The authors thank members of the Genome Research Centre for their support and technical assistance. This work is partially supported by the Edward Sai-Kim Hotung Pediatric Education and Research Fund, the Research Post-graduate Studentship (EKPK) and the Outstanding Researcher Award (YLL) of The University of Hong Kong.

Funding to pay the Open Access publication charges for this article was provided by The Shun Tak District Min Yuen Tong.

The authors have declared no conflicts of interest.


   References
Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Acknowledgements
 References
 

  1. Harper JW, Adami GR, Wei N, Keyomarsi K, Elledge SJ. (1993) The p21 Cdk-interacting protein Cip1 is a potent inhibitor of G1 cyclin-dependent kinases. Cell 75:805–16.[CrossRef][ISI][Medline]
  2. Xiong Y, Hannon GJ, Zhang H, Casso D, Kobayashi R, Beach D. (1993) p21 is a universal inhibitor of cyclin kinases. Nature 366:701–4.[CrossRef][Medline]
  3. Gartel AL, Serfas MS, Tyner AL. (1996) p21-negative regulator of the cell cycle. Proc Soc Exp Biol Med 213:138–49.[Abstract]
  4. Harada K and Ogden GR. (2000) An overview of the cell cycle arrest protein, p21 (WAF1). Oral Oncol 36:3–7.[CrossRef][ISI][Medline]
  5. Santiago-Raber ML, Lawson BR, Dummer W, et al. (2001) Role of cyclin kinase inhibitor p21 in systemic autoimmunity. J Immunol 167:4067–74.[Abstract/Free Full Text]
  6. Li R, Waga S, Hannon GJ, Beach D, Stilman B. (1994) Differential effects by the p21 CDK inhibitor on PCNA-dependent DNA replication and repair. Nature 371:534–7.[CrossRef][Medline]
  7. El-Deiry WS, Tokino T, Velculescu VE, et al. (1993) WAF1, a potential mediator of p53 tumor suppression. Cell 75:817–25.[CrossRef][ISI][Medline]
  8. Sangfelt O, Erickson S, Castro J, et al. (1999) Molecular mechanisms underlying interferon-alpha-induced G0/G1 arrest: CKI-mediated regulation of G1 Cdk-complexes and activation of pocket proteins. Oncogene 18:2798–810.[CrossRef][ISI][Medline]
  9. Hobeika AC, Etienne W, Torres BA, Johnson HM, Subramaniam PS. (1999) IFN-{gamma} induction of p21 (WAF1) is required for cell cycle inhibition and suppression of apoptosis. Interferon Cytokine Res 19:1351–61.[CrossRef]
  10. Deng C, Zhang P, Harper JW, Elledge SJ, Leder P. (1995) Mice lacking p21CIP1/WAF1 undergo normal development, but are defective in G1 checkpoint control. Cell 82:675–84.[CrossRef][ISI][Medline]
  11. McDonald ER 3rd, Wu GS, Waldman T, El-Deiry WS. (1996) Repair defect in p21 WAF1/CIP1 -/- human cancer cells. Cancer Res 56:2250–5.[Abstract/Free Full Text]
  12. Bouneaud C, Kourilsky P, Bousso P. (2000) Impact of negative selection on the T cell repertoire reactive to a self-peptide: a large fraction of T cell clones escapes clonal deletion. Immunity 13:829–40.[CrossRef][ISI][Medline]
  13. Balomenos D, Martin-Caballero J, Garcia MI, et al. (2000) The cell cycle inhibitor p21 controls T-cell proliferation and sex-linked lupus development. Nat Med 6:171–6.[CrossRef][ISI][Medline]
  14. Lawson BR, Baccala R, Song J, Croft M, Kono DH, Theofilopoulos AN. (2004) Deficiency of the cyclin kinase inhibitor p21 (WAF-1/CIP-1) promotes apoptosis of activated/memory T cells and inhibits spontaneous systemic autoimmunity. J Exp Med 16:547–57.
  15. Gaffney PM, Kearns GM, Shark KB, et al. (1998) A genome-wide search for susceptibility genes in human systemic lupus erythematosus sib-pair families. Proc Natl Acad Sci USA 95:14875–9.[Abstract/Free Full Text]
  16. Gaffney PM, Ortmann WA, Selby SA, et al. (2000) Genome screening in human systemic lupus erythematosus: results from a second Minnesota cohort and combined analyses of 187 sib-pair families. Am J Hum Genet 66:547–56.[CrossRef][ISI][Medline]
  17. Ho CY, Wong CK, Li EK, Tam LS, Lam CW. (2002) Expression of cyclin B1 and cyclin dependent kinase inhibitor p21 in lymphocytes in patients with systemic lupus erythematosus. J Rheumatol 29:2537–44.[ISI][Medline]
  18. Perlman H, Bradley K, Liu H, et al. (2003) IL-6 and matrix metalloproteinase-1 are regulated by the cyclin-dependent kinase inhibitor p21 in synovial fibroblasts. J Immunol 170:838–45.[Abstract/Free Full Text]
  19. Santiago-Raber ML, Lawson BR, Dummer W, et al. (2001) Role of cyclin kinase inhibitor p21 in systemic autoimmunity. J Immunol 167:4067–74.[Abstract/Free Full Text]
  20. Tan EM, Cohen AS, Fries JF, et al. (1982) The 1982 revised criteria for the classification of systemic lupus erythematosus. Arthritis Rheum 25:1271–7.[ISI][Medline]
  21. Hochberg MC. (1997) Updating the American College of Rheumatology revised criteria for the classification of SLE. Arthritis Rheum 40:1725.[ISI][Medline]
  22. Kong EK, Prokunina-Olsson L, Wong WH, Lau CS, Chan TM, Alarcon-Riquelme M, Lau YL. (2005) A new haplotype of PDCD1 is associated with rheumatoid arthritis in Hong Kong Chinese. Arthritis Rheum 52:1058–62.[CrossRef][ISI][Medline]
  23. Livak KJ and Schmittgen TD. (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2-delta delta C(T). Method 25:402–8.[CrossRef]
  24. Michieli P, Chedid M, Lin D, Pierce JH, Mercer WE, Givol D. (1994) Induction of WAF1/CIP1 by a p53-independent pathway. Cancer Res 54:3391–5.[Abstract/Free Full Text]
  25. Herkel J, Mimran A, Erez N, et al. (2001) Autoimmunity to the p53 protein is a feature of systemic lupus erythematosus (SLE) related to anti-DNA antibodies. J Autoimmun 17:63–9.[CrossRef][ISI][Medline]
  26. Hiyama H, Iavarone A, Reeves SA. (1998) Regulation of the cdk inhibitor p21 gene during cell cycle progression is under the control of the transcription factor E2F. Oncogene 16:1513–23.[CrossRef][ISI][Medline]
  27. Chakravarti A. (1999) Population genetics—making sense out of sequence. Nat Genet Suppl 21:56.
  28. Gordon D, Finch SJ, Nothnagel M, Ott J. (2002) Power and sample size calculations for case-control genetic association tests when errors are present: application to single nucleotide polymorphisms. Hum Hered 54:22–33.[CrossRef][ISI][Medline]
  29. Gordon D and Ott J. (2001) Assessment and management of single nucleotide polymorphism genotype errors in genetic association analysis. Pac Symp Biocomput 6:18–29.
  30. Huizinga TWJ, Pisetsky DS, Kimberly RP. (2004) Associations, populations, and the truth. Arthritis Rheum 50:2066–71.[CrossRef][ISI][Medline]
Submitted 3 March 2006; revised version accepted 11 May 2006.
Add to CiteULike CiteULike   Add to Connotea Connotea   Add to Del.icio.us Del.icio.us    What's this?



This Article
Right arrow Abstract Freely available
Right arrow FREE Full Text (PDF) Freely available
Right arrowOA All Versions of this Article:
46/2/220    most recent
kel210v1
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in ISI Web of Science
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Add to My Personal Archive
Right arrow Download to citation manager
Right arrow Search for citing articles in:
ISI Web of Science (3)
Right arrow Disclaimer
Google Scholar
Right arrow Articles by Kong, E. K.-P.
Right arrow Articles by Lau, Y.-L.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Kong, E. K.-P.
Right arrow Articles by Lau, Y.-L.
Related Collections
Right arrow Systemic Lupus Erythematosus and Autoimmunity
Right arrow Immunogenetics
Social Bookmarking
Add to CiteULike   Add to Connotea   Add to Del.icio.us  
What's this?