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p53 and reproduction

  • Hey-Joo Kang
    Correspondence
    Correspondence: Hey-Joo Kang, M.D., The Ronald O. Perelman and Claudia Cohen Center for Reproductive Medicine, Weill Cornell Medical College, 1305 York Avenue, New York, New York 10021.
    Affiliations
    The Ronald O. Perelman and Claudia Cohen Center for Reproductive Medicine, Weill Cornell Medical College, New York, New York
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  • Zev Rosenwaks
    Affiliations
    The Ronald O. Perelman and Claudia Cohen Center for Reproductive Medicine, Weill Cornell Medical College, New York, New York
    Search for articles by this author
      Tumor protein 53 (TP53) and its related family of p63 and p73 are tumor suppressor genes that regulate cellular activity to enhance longevity. p53 binds to specific response elements in DNA, modulating the transcription of genes that govern the major defenses against tumor growth. Additional members of the p53 family are involved with male and female germ cell survival. Although the majority of studies have focused on p53 as a tumor suppressor gene, little is known about its function in normal cellular processes. Polymorphisms of TP53 codon 72 that alter activity levels have been studied with respect to implantation in both the murine and human models. TP53 codon 72 (arginine) exhibits higher rates of apoptosis and leukemia inhibitory factor expression, whereas the C allele (proline) reduces leukemia inhibitory factor expression. Here, we review the role of p53 and the family of p53 proteins, along with the potential effect of p53 polymorphisms on reproduction.

      Key Words

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      Tumor protein 53 (TP53) is a tumor suppressor gene responsible for a host of protective mechanisms aimed at species survival. Commonly referred to as the “guardian of the genome,” p53 binds to specific response elements in DNA, modulating the transcription of genes that govern the major defenses against tumor growth. In response to stress signals, the p53 protein helps with genomic stability by several mechanisms. It can hold cell division at the G1/S phase, thus allowing time to activate DNA repair systems to fix the damage before continuing on with the cell cycle. If DNA damage proves to be irreparable, the p53 protein induces apoptosis or cell senescence, preventing proliferation of cancer cells. The progressive growth of cancer is reliant on neovascularization, without which tumors remain clinically benign. TP53 stimulates the production of thrombospondin-1, a potent inhibitor of angiogenesis, reducing spread of cancer cells (
      • Dameron K.M.
      • Volpert O.V.
      • Tainsky M.A.
      • Bouck N.
      The p53 tumor suppressor gene inhibits angiogenesis by stimulating the productio nof thrombospondin.
      ). Malfunction of the p53 pathway is a hallmark of human tumors: 50% of tumors harbor mutations in the p53 gene and 80% have dysfunctional p53 signaling. Located on chromosome 17 in humans, p53 is highly conserved across evolutionary time periods. The role of p53 in species survival is evidenced in individuals who carry mutations in the gene. Those who inherit only one functional copy of the TP53 gene have Li-Fraumeni syndrome, characterized by early adulthood sarcomas and breast and brain cancers. Although the majority of studies have focused on p53 as a tumor suppressor gene, little is known about its function in normal cellular processes.
      The function of p53 has been modified genetically in mice by targeted disruption of the gene. Mice homozygous for the Trp53 null allele (p53−/−) are phenotypically normal at birth but die as early as 4–6 months of age, primarily from T cell lymphomas (
      • Blackburn A.C.
      • Jerry D.J.
      Knockout and transgenic mice of Trp53: what have we learned about p53 in breast cancer?.
      ). They are also highly susceptible to developing tumors despite minimal exposure to radiation. In order to study loss of function effects, conditional knockouts in the gene are created by inserting recombination sites on both sides of the gene. A recombinase is then introduced to delete the gene segment between them, achieving tissue-specific gene deletion while allowing for individual survival of p53 null mice (
      • Donehower L.A.
      • Harvey M.
      • Slagle B.L.
      • McArthur M.J.
      • Montgomery Jr., C.A.
      • Butel J.S.
      • et al.
      Mice deficient for p53 are developmentally normal but susceptible to spontaneous tumours.
      ). This technique has facilitated the study of p53 effects on various tissue types, from breast cancer to sarcomas of the intestinal tract. When these mice do survive to breeding age, the litter sizes are approximately 25% smaller due to embryonic death from failure of anterior neural tube closure (
      • Armstrong J.F.
      • Kaufman M.H.
      • Harrison D.J.
      • Clarke A.R.
      High-frequency developmental abnormalities in p53-deficient mice. Current biology.
      ).
      The p53 family includes homologs p63 and p73. Both proteins show a 65% sequence similarity to p53 in the DNA binding domain. Studies using knockout mice demonstrate different biologic functions despite this degree of sequence homology. p63 is the most ancient member of the p53 family, and there are two main isoforms, each with distinct functions. The p63 isoform ΔNp63α plays an important role in maintaining a stem cell population in the basal layer and in the development of stratified epithelial tissue. Inactivation of this p63 isoform leads to a lack of multilayered skin, with hallmark features of cleft lip/palate syndrome, limb truncations, and mammary gland deformity (
      • Deutsch G.B.
      • Zielonka E.M.
      • Coutandin D.
      • Weber T.A.
      • Schäfer B.
      • Hannewald J.
      • et al.
      DNA damage in oocytes induces a switch of the quality control factor TAp63alpha from dimer to tetramer.
      ). The isoform TAp63α is constitutively expressed in the female germ cell, when oocytes are most vulnerable to DNA damage while arrested in a tetraploid state during the diplotene stage (dictyate phase) of meiosis I. It serves as a quality control factor to ensure oocytes with DNA damage are eliminated by apoptosis before they can be recruited for ovulation, and makes them less sensitive to DNA damage (
      • Suh E.K.
      • Yang A.
      • Kettenbach A.
      • Bamberger C.
      • Michaelis A.H.
      • Zhu Z.
      • et al.
      p63 protects the female germ line during meiotic arrest.
      ). Knockout mice for TAp63α display normal oogenesis and folliculogenesis, but exposure to minimal degrees of radiation (0.45 Gy) results in a dramatic loss of primordial follicles within five days.
      The homolog P73 serves a unique role in both oogenesis and spermatogenesis. Male and female TAp73−/− mice are both infertile despite normal mating behavior. In females, infertility results from ovulated oocytes becoming trapped beneath the ovarian bursa, unable to transfer to the fallopian tube. Upon closer examination, these oocytes also show a high degree of spindle abnormalities, such as multipolar spindles, and spindle relaxation and scattering (
      • Gebel J.
      • Tuppi M.
      • Krauskopf K.
      • Coutandin D.
      • Pitzius S.
      • Kehrloesser S.
      • et al.
      Control mechanisms in germ cells mediated by p53 family proteins.
      ). TP73 also ensures correct chromosomal segregation (
      • Tomasini R.
      • Tsuchihara K.
      • Tsuda C.
      • Lau S.K.
      • Wilhelm M.
      • Ruffini A.
      • et al.
      TAp73 regulates the spindle assembly checkpoint by modulating BubR1 activity.
      ) in female mouse oocytes, and its concentration decreases with female age. If this holds true in humans, the waning presence of p73 over time may contribute to increased abnormal chromosomal segregation with maternal age.
      As sperm mature, the spermatogonia migrate from the basement membrane and through the blood–testis barrier to reach the lumen of the seminiferous tubule. The occluding junctions of the Sertoli cell form the blood–testis barrier and partition the interstitial blood compartment of the testis from the adluminal compartment of the seminiferous tubules. Sertoli cells control the entry and exit of nutrients and hormones to regulate the internal environment, and provide protection from noxious elements in the circulation. They also provide multiple levels on which the developing sperm can dock, as well as structural and metabolic support to the maturing spermatocyte. TP73 appears to balance the expression of proteases, protease inhibitors, and integrins to allow for a functional blood–testis barrier (
      • Beumer T.L.
      • Roepers-Gajadien H.L.
      • Gademan I.S.
      • van Buul P.P.
      • Gil-Gomez G.
      • Rutgers D.H.
      • et al.
      The role of the tumor suppressor p53 in spermatogenesis.
      ). TAp73−/− male mice are infertile due to defective cell–cell adhesions between developing germ cells and Sertoli cells. Because of this loss of contact, histochemical analysis of seminiferous tubules of TAp73−/− mice shows a dramatic loss of maturing germ cells and spermatozoa (
      • Gebel J.
      • Tuppi M.
      • Krauskopf K.
      • Coutandin D.
      • Pitzius S.
      • Kehrloesser S.
      • et al.
      Control mechanisms in germ cells mediated by p53 family proteins.
      ). Without the tight contact between the maturing germ cell and Sertoli cells, development of sperm cells is severely diminished.

      Polymorphisms of p53

      A single nucleotide polymorphism (SNP) is a germline mutation within a gene, often at a specific nucleotide base. The majority of polymorphisms are intronic and phenotypically silent without biologic consequence; however, single base substitutions in exons can cause gain-of-function or loss-of-function changes. TP53 is unique among tumor suppressor genes in that a host of missense mutations can occur, generating a wide range of p53 proteins with varying levels of activity. The TP53 codon 72 polymorphism (rs 1042522) is one such mutation. The wild-type C allele and variant G allele lead to a proline or arginine in codon 72, respectively. The G allele has a higher propensity to induce apoptosis and protect cells from neoplastic development, whereas the C allele has weaker p53 apoptotic activity. A potent tumor suppressor gene like p53 needs a negative regulator to inhibit apoptosis in unstressed conditions. Mouse double minute 2 homolog (MDM2) is the key negative feedback regulator of TP53 activity. An important polymorphism in the MDM2 gene is located at nucleotide 309. This T→G change increases MDM2 transcription and attenuates TP53-mediated apoptosis. The T allele is the wild-type allele, but a high frequency of the G allele suggests it is under positive evolutionary selection—despite posing a higher risk for cancer. This suggests that stronger MDM2 activity is protective in some way by keeping p53 or an associated gene product under control.
      At the start of implantation, blastocysts are in close contact with the luminal epithelium. This is followed by apoptosis of the luminal epithelium, allowing trophoblast cells to migrate into the underlying endometrial stroma. The stroma responds with rapid proliferation and differentiation to form the decidua. Implantation is also dependent on establishing normal placental vasculature, coordinating vascular endothelial cell–specific growth factors with an exchange of signals between these cells (
      • Red-Horse K.
      • Zhou Y.
      • Genbacev O.
      • Prakobphol A.
      • Foulk R.
      • McMaster M.
      • et al.
      Trophoblast differentiation during embryo implantation and formation of the maternal-fetal interface.
      ). If implantation does not occur, the uterus becomes non-receptive and re-enters the menstrual cycle. Many of the steps involved in implantation—apoptosis, angiogenesis, maintaining genomic stability—are also functions regulated by p53. Thus, it would be logical to assume that the p53 gene, critical to survival and longevity of an individual, could also play a broader role in species survival by facilitating implantation efficiency.
      The importance of leukemia inhibitory factor (LIF) in implantation was demonstrated in the murine model, where female mice carrying a null mutation in the LIF gene were sterile due to a complete failure to implant blastocysts. Replacement of estradiol and progesterone was not sufficient to induce decidualization in LIF-deficient mice. In ovariectomized mice, injections of LIF at doses as small as 25 ng permitted decidualization and blastocyst implantation with normal embryogenesis. Intraperitoneal injection of LIF into pregnant LIF-deficient females was sufficient to rescue embryo implantation and allow for LIF-deficient females to give birth to viable offspring. In aggregate, LIF is essential for initiating implantation but not required for embryo development or the maintenance of pregnancy (
      • Chen J.R.
      • Cheng J.G.
      • Shatzer T.
      • Sewell L.
      • Hernandez L.
      • Stewart C.L.
      Leukemia inhibitory factor can substitute for nidatory estrogen and is essential to inducing a receptive uterus for implantation but is not essential for subsequent embryogenesis.
      ).
      LIF was identified as a p53 target gene in a study by Hu et al. in 2007 (
      • Hu W.
      • Feng Z.
      • Teresky A.K.
      • Levine A.J.
      p53 regulates maternal reproduction through LIF.
      ). A putative DNA response element was identified within intron 1 of both mouse and human LIF genes, and it was shown to regulate both basal and inducible transcription of LIF. Lower LIF levels were measured in the uteri of p53 null mice when compared to wild-type mice on day 4 of pregnancy, when higher LIF levels are critical to permit implantation. Subsequently, litter sizes in p53 null mice with lower LIF levels were significantly smaller than in wild-type mice, with full recovery of litter sizes after intraperitoneal injection of LIF. Although 70% of the litter was born healthy, the remainder had developmental abnormalities of the neural tubes, suggesting that exogenous delivery of LIF was not enough to completely restore natural conditions for implantation (
      • Hu W.
      • Feng Z.
      • Teresky A.K.
      • Levine A.J.
      p53 regulates maternal reproduction through LIF.
      ). Polymorphisms of TP53 codon 72 that alter activity levels have been studied with respect to implantation in both the murine and human models. TP53 codon 72 with arginine exhibits higher rates of apoptosis and LIF expression, whereas the C allele reduces LIF expression.

      Polymorphisms of p53 codon 72 and miscarriage

      Since p53 induces LIF expression, it is logical to consider whether patients with recurrent miscarriages or infertility have a higher rate of p53 polymorphisms than fertile controls. The proline-rich domain at codon 72 is associated with reduced LIF expression, lower rates of apoptosis, and higher G1 cell cycle arrest than arginine at codon 72. In inducible cell lines with equal levels of p53 and in tumor cell lines with endogenous wild-type p53, the Arg72 form of p53 is at least five times better at inducing apoptosis compared to the Pro72 variant. To date, there have been seven studies examining the role of p53 in unexplained recurrent miscarriage.
      The first study in 2005 compared 175 Caucasian women with at least three spontaneous, consecutive miscarriages before 20 weeks gestation to 143 fertile controls with at least one live birth and no history of miscarriage. The homozygous Pro/Pro genotype was found more often in women with recurrent miscarriage compared to fertile controls (12.6% vs. 7%), respectively. Those who are heterozygous for the Pro/Arg genotype were also found more frequently in the recurrent miscarriage group compared to controls, and the presence of a proline allele was found in 32.6% of recurrent loss patients compared to 24.5% of the control population (P=.03). The authors theorized this difference may be caused by impaired placental structure and differentiation to provide adequate gas and nutrient exchange between the mother and the fetus due to prolonged G1 cell cycle arrest. Similarly, reduced apoptosis in carriers of the Pro allele may lead to unhealthy tissue permitted to continue on and populate the placenta, reducing efficiency to transport nutrients (
      • Pietrowski D.
      • Bettendorf H.
      • Riener E.K.
      • Keck C.
      • Hefler L.A.
      • Huber J.C.
      • et al.
      Recurrent pregnancy failure is associated with a polymorphism in the p53 tumour suppressor gene.
      ). A follow-up study by a different group in 2006 examined women with recurrent implantation failure (RIF) as well as recurrent pregnancy loss (RPL). Recurrent implantation was defined in this study as a cumulative of eight cleaved embryos or four blastocysts transferred with HCG concentrations of <5 mIU/mL. Recurrent miscarriage was defined as two or more consecutive spontaneous abortions with no known etiology. There were 70 RIF, 205 RPL, and 20 fertile controls compared in this study. The Pro/Pro genotype was found in 7%, 4%, and 0%, respectively, and at least one Pro allele was found in 30%, 18%, and 19%, respectively (P=.003). While the incidence of the Arg genotype was noted to be reduced in RIF patients compared to all other study patients, the Pro genotype was found to be higher in RIF patients. The p53 gene is also a potent inducer of angiogenesis via the hypoxia-inducible factor 1α and vascular endothelial growth factor pathways. Since the activity of p53 is reduced with the proline variant allele, the overall effect would be reduced rates of apoptosis with higher rates of cell cycle arrest, perhaps leading to reduced angiogenesis at the site of implantation (
      • Kay C.
      • Jeyendran R.S.
      • Coulam C.B.
      p53 tumour suppressor gene polymorphism is associated with recurrent implantation failure.
      ).
      Under normal, unstressed conditions, MDM2 binds to the transactivation domain of p53 and ubiquitylates the protein, targeting it for degradation. These proteins are balanced through a negative feedback loop, which is reversed by an increase in p53 levels when a stress signal is received. An SNP in the MDM2 protein at location 309 from thymine to guanine enhances transcription of MDM2, reducing p53-mediated apoptosis. The first study to examine MDM2 SNP 309 evaluated allele frequency in women with idiopathic recurrent miscarriages. Among 95 women experiencing recurrent miscarriages, the MDM2 G/G genotype was found in 32.6% compared to 18.3% in 164 fertile controls. A particular strength of this study was that the average age of patients was 28, reducing the incidence of genetic aneuploidy as a cause for miscarriage. For women who underwent a dilatation and curettage, the villous samples were also genotyped. The G/G genotype was found in 28.3% versus 12.5% (P<.05), suggesting increased MDM2 activity, thus reduced p53 activity, in villi of women suffering from recurrent miscarriages (
      • Fang Y.
      • Kong B.
      • Yang Q.
      • Ma D.
      • Qu X.
      MDM2 309 polymorphism is associated with missed abortion.
      ). A follow-up study by the same group genotyped both p53 codon 72 and MDM2 SNP 309 in 60 women with recurrent miscarriage and compared them to genotypes of 64 women with at least one live birth and no history of miscarriage. There was no difference in the p53 codon 72 proline versus arginine genotypes between the recurrent loss and control groups; however, the recurrent loss group had a higher incidence of the G/G genotype: 28.3% versus 12.5% (P<.05) (
      • Fang Y.
      • Kong B.
      • Yang Q.
      • Ma D.
      • Qu X.
      The p53-HDM2 gene-gene polymorphism interaction is associated with the development of missed abortion.
      ). This was followed by a case-control study in Brazil, comparing 120 women with two or more pregnancy losses with 143 fertile controls with no history of pregnancy loss. After correcting for smoking, alcohol consumption, ethnicity, and number of previous pregnancies, p53 Arg/Arg and MDM2 T/T genotypes were found to increase the risk of RPL (OR = 2.58; CI = 1.31–5.07). The estimated probability for miscarriage was 71% for carriers and 48% for non-carriers. However, this difference no longer existed when evaluating a subgroup of African ancestry, and only held for women reporting European ethnicity. The ethnic heterogeneity of this population compared to previous studies may have contributed to the difference in allelic frequency between the study and control population (
      • Fraga L.R.
      • Dutra C.G.
      • Boquett J.A.
      • Vianna F.S.
      • Gonçalves R.O.
      • Paskulin D.D.
      • et al.
      p53 signaling pathway polymorphisms associated to recurrent pregnancy loss.
      ). The majority of studies include homogenous populations, as in the case of an Iranian study published in 2011 (
      • Firouzabadi R.D.
      • Ghasemi N.
      • Rozbahani M.A.
      • Tabibnejad N.
      Association of p53 polymorphism with ICSI/IVF failure and recurrent pregnancy loss.
      ) comparing allelic frequencies of RPL, RIF, and fertile control patients. Their results, although hindered by small sample sizes, supported earlier studies showing higher rates of pro/pro genotype in women with RPL. The average age in this study was 29, which again reduced the risk of aneuploidy as a confounding factor for this group of women with recurrent losses. A meta-analysis incorporating several of these studies also found PRL to be associated with the pro/pro polymorphism, most notably in Caucasians (
      • Chen H.
      • Yang X.
      • Wang Z.
      Association between p53 Arg72Pro polymorphism and recurrent pregnancy loss: an updated systematic review and meta-analysis.
      ). As a group, the studies genotyping p53 and MDM2 suggest a higher rate of proline at codon 72 in women with RPL, theorized to be from reduced apoptotic activity, diminished angiogenesis, longer cell cycle arrest, or an interplay of these various p53 functions.

      Polymorphisms of p53 codon 72 and in vitro fertilization outcomes

      Women with unexplained infertility became the next focal point of study in p53 research. Because age-related chromosomal aneuploidy is the major underlying cause for infertility, the next study included only infertile women <35 years old using autologous oocytes or older women using donor oocytes. A total of 272 women were included and compared to women in the general population. The frequency of p72, which is weaker in the induction of LIF expression, was enriched in the in vitro fertilization (IVF) group (33.1% vs. 22.7%; P<.005) compared to the control population. The enrichment of p72 proline was more significant in women <35, who are more likely to have implantation problems compared with older women whose infertility is more often from aneuploidy. The study went further to investigate implantation and pregnancy rates of women with the p53 Pro72 allele compared to the wild-type arginine. Implantation rates were defined as sacs/embryo transferred, and clinical pregnancy considered as fetal cardiac activity at 6–7 weeks of gestation. The implantation rate was lower in women homozygous for Pro72 (19%) compared with patients carrying at least one allele of Arg72 (19% vs. 42%; P=.0028), as were the clinical pregnancy rates (38% vs. 61%) (
      • Kang H.J.
      • Feng Z.
      • Sun Y.
      • Atwal G.
      • Murphy M.E.
      • Rebbeck T.R.
      • et al.
      Single-nucleotide polymorphisms in the p53 pathway regulate fertility in humans.
      ).
      A study published the same year compared infertile women who conceived on their first IVF attempt with those whose first cycle failed and who continued on to subsequent IVF cycles. There were no exclusion criteria; thus, the study included 1,056 patients with an average age of 35–36. They found no difference in the G and C allele frequency; however, the authors were comparing infertile women who implanted with their first transfer to other infertile women who implanted in subsequent cycles, rather than fertile controls. In addition, the average age was 6–8 years older than in previous studies, permitting a higher rate of aneuploidy as a cause of implantation failure. The lack of exclusion criteria may have confounded their results as well, allowing for male factor cases and women with secondary infertility to be included in the study (
      • Patounakis G.
      • Treff N.
      • Tao X.
      • Lonczak A.
      • Scott Jr., R.T.
      • Frattarelli J.L.
      The p53 codon 72 single nucleotide polymorphism lacks a significant effect on implantation rate in fresh in vitro fertilization cycles: an analysis of 1,056 patients.
      ). Interestingly, the patient cohort in this study had a p53 P72 allele frequency (33%) very similar to the patient cohort in previous studies (33.1%). Finally, a recent study of 1,450 first-time IVF patients with an average age of 30–31 examined both p53 and MDM2 polymorphisms. When comparing allele frequencies and IVF outcome, the authors found a higher frequency of the C allele in the group with a positive clinical pregnancy (52.1% vs. 47.4%). However, the pregnancy rate overall was only 39% (569/1,449), lower than average in this patient demographic (
      • Chan Y.
      • Zhu B.
      • Jiang H.
      • Zhang J.
      • Luo Y.
      • Tang W.
      Influence of TP53 Codon 72 Polymorphism alone or in combination with HDM2 SNP309 on human infertility and IVF outcome.
      ). Collectively, the conflicting results may be explained by genetic heterogeneity, as different distributions for the p53 polymorphism vary by regional or ethnic groups. Groups that include women with advanced maternal age or have a small sample size may miss the subtle role in human fertility that SNPs may play.

      Conclusion

      p53 has evolutionary conserved functions besides tumor suppression, as target transcription factors are maintained from invertebrates to vertebrates. Indeed, the existence of p53-like proteins in short-lived organisms from fireflies to worms that do not exhibit adult cancers implies that tumor suppression was not the original purpose of p53 (
      • Hu W.
      • Feng Z.
      • Teresky A.K.
      • Levine A.J.
      p53 regulates maternal reproduction through LIF.
      ). Thus, it is feasible that the function of p53 and its family of proteins that evolved in the human as a tumor suppressor can also extend to other functions such as reproduction.
      Although the majority of implantation failures and recurrent miscarriages are due to embryonic chromosomal aneuploidy, there remains a subset of euploid embryos that fail to implant. Some of these failed implantation/early miscarriages from PGS-tested embryos are attributable to mosaicism, but there is also a portion that fail due to an as yet unknown uterine factor. p53 may function as a post-zygotic selection step to inhibit proliferation of embryonic and trophoblastic cells. Furthermore, investigations have shown that an abundance of p53 protein is produced by the human placenta in abnormal pregnancies; thus, p53 is suspected as an important factor for the pathogenesis of placental disorders through the induction of trophoblastic apoptosis (
      • Hu C.
      • Smith S.D.
      • Pang L.
      • Sadovsky Y.
      • Nelson D.M.
      Enhanced basal apoptosis in cultured term human cytotrophoblasts is associated with a higher expression and physical interaction of p53 and Bak.
      ).
      Elucidating the impact of p53 on reproduction is difficult. Unlike germline mutations that create obvious disease patterns in Li-Fraumeni syndrome, polymorphisms are thought to impact the host in a modest fashion, such as causing earlier onset of disease or a small increase in the risk of cancer or subtle diminishment in fertility. These small differences present a challenge in verifying the biological impact of polymorphisms on fertility. Case-control studies comprise the majority of investigative work so far and have not reached the scope or size to be conclusive. To reach a definitive answer of the impact of p53 on reproduction, large population-based studies will be required. In addition, there remain several unanswered questions. Perhaps the variance in results is due to our focus on p53 polymorphism solely in the mother, rather than in the blastocyst. If the p53 activity level in the blastocyst contributes to implantation success, might testing the blastocyst itself or the male partners for p53 polymorphisms be a more comprehensive method of study? Further work in this area will help to define these relationships. Potential therapeutic options would range from adding LIF during the implantation period to selectively modifying the polymorphism to adjust the degree of p53 activity. Any modifications to p53 activity would be done with caution, as the cellular activity of p53 has not been fully elucidated, and the goal of increasing reproductive efficiency may come with unintended consequences, from malformations of the offspring to reduced longevity. Ultimately, the selected alleles in the p53 pathway discussed here for both fertility and cancer risk may demonstrate a cooperative pleiotropy, whereby a single gene influences multiple phenotypic traits with a common goal (
      • Hu W.
      • Feng Z.
      • Teresky A.K.
      • Levine A.J.
      p53 regulates maternal reproduction through LIF.
      ). As we continue our study of this fascinating gene, cancer protection and efficient reproduction appear to be merely two of the many protective mechanisms regulated by the p53 family of proteins.

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