The Fragile X Syndrome

The Fragile X SyndromeThe soft X syndrome is unity of the close prevalent psychic maladroitness problems that be inherited to ingredientrations. The clinical features of people with this syndrome be very subtle and hard to diagnose. Recent come along in the field of molecular biology and ingredienttics has bylined the molecular magnetic variation, that causes the syndrome was a triplet resort mutation. Due to repetitive CGG repeat the respective proteins are non expressed. New molecular methods including direct investigating analysis and PCR analysis, slang simplified the process of diagnosis. The nature of the gene, their respective gene product and its function has not been yet found clearly. heritage may be found overdue to common ancestral fuse of X chromosome at birth.IntroductionFragile S syndrome is a genetical disorder that affects the intellectual, physical and moral factors of a benignant being. It is also k flatn to be martin-bells syndrome and the effects range widely from mild to severe. It is caused by the mutation on the X chromosome of the individualistic in a single gene called the Fragile X noetic retardation gene (FMR1). Psychological problems such as mental retardation may be caused by both main factors, the physical environment or the genetic factor that is hereditary. As we chiefly focus on the genetic factor, the FRAXA locus in xq27.3 is associated in ca exploitation fragile X mental retardation. therefore based on molecular genetic testing of FMR1 gene, fragile X syndrome plenty be diagnosed. Women are hardly 50% abnormal by this syndrome when compared to males due to the fact that they select two X chromosomes where as males have one. Most common symptoms found are seizures, mood instability, attention deficit, sensory oer stimulation, aggression, autism, speech disorder and sleep differences. This syndrome also shows certain physical symptoms such as long constringe face, large ears enlarged testacles in males, flexible joints etc., various researches is being done across the world to detect a solution for the disease.Various genetic techniques has been developed and handled to identify the fragility of the chromosome. There has been so much improvement in research but a definite solution has not been obtained. The treatment for this syndrome is unremarkably a multidisciplinary approach which includes occupational therapy, medical managements, education and linguistics.The Fragile X MutationGenes are the precursors of specialized protein molecules which in turn are particular for various functions of the body. The major cause for the fragile x syndrome is due to the mutation in one single gene called the fragile x mental retardation 1 (FMR1) gene which is the precursor for the fragile x mental retardation protein FMRP. This protein is responsible for the chemical formula development and functioning of the brain. Men have wholly one x chromosome so presence of mutation in that chromosome volition cause the disease whereas in womanishs, they have two x chromosomes and hence full mutation in one copy makes them carriers of the syndrome and they may be affected partially according to the totality of mutation and reduce of carrells expressing the FMR-1 gene copy. The inheritance found on the chromosomes is termed as x linked recessive inheritance which is more complex than the normal x linked genes.Fig.1 X chromosome with fragile site 1 Fig. 2 A dart of X chromosomes showing a fragile site from both a male and a feminine 2The number of CGG repeats on the FMR-1 gene studys the complexity of the syndrome. The repeats are classified as curtly, medium and long repeats. The short repeat of about six to fifty times which is found very common. These short repeats are broadly unstable and do not definitely cause the syndrome. Yet a genetic counseling along with certain tests is recommended. The medium repeat is about 50 to 200 times and is called pe rmutation.The fragile x mental retardation protein (FMRP) has lower risk of the syndrome as short sequence repeats. The long repeats are usually more than 200 and are termed as full mutation where the complete FMR-1 gene is altered and production of the FMRP protein is completely stopped. Among the people with full mutation or long repeats, men will have the fragile x syndrome and women will be carriers.Fragile x mental retardation protein (FMRP)The FMRP protein is found in the ribo-nucleoprotein complex and is encoded by the FMR1 gene. The FMRP weighs up to 60-70 kD. This protein is associated with the polyribosome or polysomes. Two RNA-binding domains, KH domains or K homology domains are possessed by this protein and it binds to fetal human brain to 4% approximately. It also has the ability to bind to its own mRNA. Even a clear amount of mutation in one of the KH domain could stop its interaction with the polysomes leading to the fragile x syndrome.Inheritance of fragile x synd romeMales have xy chromosome and hence have only one FMR-1 gene where as distaffs have xx chromosomes and hence they have two FMR-1 genes. On the F1 generation for each one parent transfers one chromosome each to the offspring where the transfer of the FMR-1 gene is determined. Therefore the possibilities of their offsprings being affected are grouped to a lower place two conditions.Condition-1 3If a male has a mutated gene in his chromosome it can be transferred only to his daughter because only the Y chromosome can be transferred to his son by him. So if he was crossed with a female with normal genes all their sons will be normal and the daughters will have one fragile gene and remain as carriers.fatherymotherxx (carrier daughter)Xy (normal son)xx (carrier daughter)Xy (normal son) fragile x geneCondition-2 4If a female has one mutated gene in her chromosome and is crossed with a normal male then there is 50% chance of all the offsprings, be it male or female to have the syndro me.fatherxymotherx (carrier daughter) y (fragile son)xxx(normal daughter)Xy (normal son) fragile x geneSymptomsThe symptoms of fragile x syndrome are categorized into PhysicalLarge nerve centersProminent eyebrowLarge testeclesSeizuresCognitive developmentSocial and emotionalhyperactivityBehavioralShynessSocial anxiety spoken communication and languageRapid and repetitiveInability to adopt wordsOver-talkativeAutismFlapping of handsPoor eye contactAll these symptoms need not be necessarily seen. A combination of various symptoms may vary from person to person based on the amount of gene altered in their chromosome. sometimes there may also be no visible symptoms making the chances of early diagnosis regular worse.Molecular diagnosis 5The chromosome associated with the syndrome is classified into three major types based on the number of CGG repeats as conventionalism allelomorphthe CGG repeat in FMR1 is 6-50PCR analysis is sufficient to study all normal type genes particular point mutations and mosaicism must be studied development specific types of pcr or other molecular methods visualization is achieved either by radio-active labeling or auto radiography followed by automated sequencingAgarose gel electrophoreses may be used for simple separation analysis with stains such as ethidium bromide. Appropriate size markers and size controls are very important.Controls used for analysis mut approximately contain 50 repeats.Pre mutationThe CGG repeat in FMR1 is 55-200PCR analysis is not possible hence gray billet is al counsels preferred.Since both premutation and full mutation have methylation status, specific methylation sensitive enzymes such as EagI or NreI is used to resolve the size of the fragment.Methylated alleles are cut only by one enzyme where as non methylated normal alleles are cut by both the enzymes.Prenatal diagnosis is very important for pre mutation carriers.Rather than normal PCR a radioactive PCR can be used to test for premutation and then the result can by confirmed using southerly analysis.Full mutationThe CGG repeats in FMR1 ranges from 200 to thousandsThis can be analysed only by a southerly blot technique.At complications, if a confirmable result could not be obtained from a southern analysis then a radio active PCR can be run combined with a linkage analysis and the result can be confirmed with southern blotting.Intermediate allelesThe CGG repeats in FMR1 gene is usually between 45-55Since they are in the overlapping region between stable normal allele and unstable premutation alleles, diagnosis and interpretation is very difficultDiagnostic Tools and MethodsWith the advancement in technology desoxyribonucleic acid tests are always effective in diagnosis of fragile x syndrome. With the findings of Sutherland et al. that folic acid deficient cell culture medium was able to induce a fragile site at xg27.3 cytogenetics was the major way to determine the presence of the syndrome but after cloning of the FMR-1 gene direct methods for identifying the x linked gene has become possible. By using monoclonal antibodies specific to FMRP it is also possible to show the expression of FMR-1.The most common methods used for diagnosis in the genetic level arePolymerase Chain Reaction gray BlottingAntibody testDenaturing gradient gel electrophoresisSingle strand confirmation polymorphismNon-radioactive molecular diagnosis.Polymerase cosmic string reactionPolymerase chain reaction may be defined as a technique where one copy of a DNA is amplified into numerous copies at a rate of 2n where n is the number of cycles. It is achieved under specific conditions of temperature, along with polymerase enzyme.PCR amplification is one of the preliminary methods in diagnosing fragile x syndrome. Since the syndrome is associated with CGG repeats PCR is not considered as the best method always, since the amplification across C-G composition could be unreliable for PCR technique. However now its very much possible for a PCR to identify CGG repeats in combination with various techniques.methylation specific PCR of the FMR1 locusfluorescent methylation specific PCRmethyl-CpG-binding PCRSome of the major advantages of PCR are that it is less time consuming, a very small amount of the sample is enough to produce numerous copy and the tri-nucleotide repeat in the FMR-1 gene is accurately sized. There are also various disadvantages of this technique. When there are more than hundreds of tandem repeats it is impossible for the PCR to determine the complete mutation which may give a different result. Due to differential amplification PCR is incompetent of detecting mosacism between pre mutation and normal alleles.Fig 3 fragile x analysis using PCR 6Southern BlottingSouthern blotting is one of the best methods of diagnosing fragile x syndrome. It is modtly used as the validatory test after PCR. The variations between the mutations and permutations along with the amount of methylation occurred can be clea rly obtained by the southern blotting technique. The process can be summed up in two simple stepsStep 1 the patients DNA is digested using restriction enzymes.Step 2 southern hybridization is carried out along with specific radioactive probes after separation of FMRI region.Using southern Blotting, the differences in full mutation and pre mutation can be easily identified. Full mutations usually cause smearing of the band and are always unstable. The only advantage of this technique is that its accuracy whereas its labor intensive, time consuming. The major drawback of this method is its inability to determine the aim number of tandem repeats of the CGG nucleotides which is very much necessary in determining whether the patient is completely affected or a carrier.Fig 4 fragile x analysis by southern blot 7N refers to normalSpecific tools for analysisDNA probeA DNA probe can be defined as a single strand of DNA which act as a template to identify the target DNA molecule. To identify the fragility of chromosome on the DNA specific probes were knowing which increases the accuracy rate of the diagnosis. Chemicon (Millipore) 8 has designed a special probe named The CHEMI probe which is labeled with dioxegenin to detect the CGG repeats in the FMR-1 gene.MarkersThere were special markers called the microsatelite markers used in linkage analysis. This came to an end with the advent of modern techniques. However these markers are now being used under special circumstances like prenatal diagnosis where southern blotting has failed. Some of the markers used are DXS548, FRAXAC1 and FRAXAC2 combined with PCR. They are considerably accurate and they undergo low recombination mechanics with CGG repeats.TreatmentsThere are no gene therapies or genetic treatments ready(prenominal) for fragile x syndrome though a lot of other therapies are available which include speech-language therapy, occupational therapy, physiotherapy and behavioral therapy.There are also a large number of medications available as listed in the table infraSymptomsMedicationsSeizuresMood instability Carbamazepine (Tegretol) Valproic acid or divalproex (Depakote) Lithium carbonate Gabapentin (Neurontin) Lamotrigine (Lamictal) Topiramate (Topomax), tiagabine (Gabitril), and vigabatrin (Sabril) Phenobarbital and primidone (Mysoline) Phenytoin (Dilantin) solicitude deficit (With or without hyperactivity) Methylphenidate (Ritalin, Concerta) and dexamethamphetamine (Adderall, Dexedrine) L-acetylcarnitine Venlafaxine (Effexor) and nefazodone (Serzone) Amantadine (Symmetrel) Folic acidHyperarousalSensory over-stimulation (Often occurs with ADD/ADHD) Clonidine (Catapres TTS patches) Guanfacine (Tenex)Aggressionintermittent explosive disorderObsessive-compulsive disorder(Often occurs with anxiety and/or depression) Fluoxetine (Prozac) Sertraline (Zoloft) and citalopram (Celexa) Paroxetine (Paxil) Fluvoxamine (Luvox) Risperidone (Risperdal) Quetiapine (Seroquel) Olanzepine (Zyprexa )Sleep di sturbances Trazadone Melatonin hold over 2 symptoms and medications for FRAXA 9(*these prescriptions have serious effects. DO NOT INTAKE ANY OF THESE WITHOUT CONSULTING A MEDICAL PRACTITIONER) rate of flow ResearchGene Therapy studies are carried out on the recombination strategy of the target gene, whether removal or stand-in of the defective gene with a recombinant gene would eliminate the syndrome.Protein Replacement Therapy research is being carried out on the possibility of producing FMR protein and supplying to the patients through external sources like food or tablets.Psychopharmacology research is being carried out in finding medications for all the symptoms of fragile x syndrome.ConclusionFragile x syndrome is one of the genetic diseases that causes psychological problems due to the lack of FMR protein responsible for the mental behavior of the person. The protein is not expressed in the individual due to fragility of the FMR1 gene in the x chromosome. Though PCR and sout hern blotting are the only tools available for diagnosis they are considerably accurate and research is being carried out on various re-combinative tools for diagnosis. A complete cure has not been still devised for the syndrome though various behavioral and physical therapies help the patients get into psychological strength.