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Molecular Basis of Disease—Pathology, Laboratory Medicine - Part 1 (20 test)
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- The most common mutation causing cystic fibrosis (CF-MIM*219700) in Caucasians is ΔF508, accounting for 70% of mutations—ΔF508 refers to deletion of a phenylalanine residue (F in single-letter amino acid code) at the 508th amino acid of the cystic fibrosis transmembrane regulator (CFTR) protein. Which of the following would best describe DNA diagnosis of cystic fibrosis patients who are homozygous for the ΔF508 mutation?
- Polymerase chain reaction using primers surrounding nucleotide #1522 of the CF DNA coding sequence positive hybridization with oligonucleotide specific for the ΔF508 allele
- Northern blotting of sputum RNA samples developed with CFTR gene probes, yielding two abnormally sized RNA transcripts
- Southern blotting using restriction endonucleases that cleave at each end of the CFTR coding sequence
- Western blotting of sputum proteins developed with antibody to CFTR protein, yielding two abnormally sized bands
- Polymerase chain reaction using primers that amplify a fragment surrounding nucleotide #508 of CF coding sequence followed by detection of two fragment sizes by agarose gel electrophoresis
2. A research project examines multiple chromosome regions in Italian families selected because several individuals are affected with insulin-dependent diabetes mellitus (IDDM, MIM*222100). The results reveal a single nucleotide change from A to G in a noncoding region of chromosome 10 that is present in 9 of 10 diabetics and only 6 of 30 The conclusion is best described by which of the following statements?
- Mutations in a gene on chromosome 10 cause diabetes in Italians, allowing DNA diagnosis by Southern blot with a 90% sensitivity.
- Mutations in a gene on chromosome 10 cause diabetes in Italians, allowing DNA diagnosis by PCR amplification and DNA sequencing with a 60% positive predictive value.
- A single nucleotide polymorphism on chromosome 10, detected by PCR amplification and DNA sequencing, is associated with IDDM in Italian families (sensitivity is 90%).
- A single nucleotide polymorphism on chromosome 10, detected by PCR amplification and DNA sequencing, is now available for worldwide screening of diabetes susceptibility.
- Association of a chromosome 10 gene mutation with IDDM in Italian families suggests that HLA loci may exist outside of chromosome 6.
3. Sickle cell anemia (MIM*141900) is caused by a specific mutation in the gene for β-globin, one of the two globin proteins that complex with heme to form The sickle cell anemia mutation is known to produce a single amino acid change, glutamic acid to valine, at position 6 of the β-globin peptide chain. Which of the following is the most likely mechanism for this mutation?
- Crossing over
- Two-base insertion
- Three-base deletion
- Single-base insertion
- Single-base substitution (point mutation)
4. A 2-year-old girl is the product of a normal pregnancy and delivery and developed normally until age 12 months. After learning many words, crawling, and walking, she becomes progressively unsteady and stops walking, also refusing to She begins wringing her hands frequently, has staring spells, and seems to be regressing in performance and memory. Her physician suspects a disorder called Rett syndrome (MIM*312750), and orders a diagnostic test that is based on increased DNA methylation around the MECP gene on the X chromosome. Which of the results below would be consistent with a positive diagnosis?
- Increased expression of the MECP protein
- Increased size of MECP mRNA
- Loss of methylated sites as detected by methylation-sensitive restriction endonucleases
- Gain of methylated sites as detected by methylation-sensitive restriction endonucleases
- Amplification of triplet repeats surrounding the MECP gene promoter region
5. A 6-month-old Hispanic girl has had six episodes of pneumonia, both viral and bacterial, and has spent only a few weeks outside of the Review of neonatal x-rays shows a very small thymus, and study of white cell markers suggests that both B (bursa-derived) and T (thymus-derived) cells are deficient. The autosomal recessive severe combined immune deficiency due to adenosine deaminase deficiency (ADA) (MIM*102700) is suspected and the physician-researcher team decides to try gene therapy as pioneered at other centers. Which of the following steps in performing gene therapy will be most challenging for the team?
- Cloning of the ADA gene by digestion of human genomic and retroviral DNA with the appropriate restriction endonuclease and constructing a library of recombinant molecules
- Transformation of the child’s bone marrow cells with DNA from a recombinant retrovirus that contains the ADA gene
- Engineering ADA gene insertion such that the retroviral ADA gene is replicated and regulated similarly to natural ADA genes
- Measuring ADA gene expression in transformed bone marrow cells to ensure sufficient immune function after transplant
- Suppression of the child’s immune system so that transplanted bone marrow cells with recombinant ADA genes will not be rejected
6. A 9-month-old girl presents to hematology clinic after her 9-month checkup showed Her serum iron and transferrin levels were normal, and her hemoglobin electrophoresis demonstrated an abnormal hemoglobin in addition to A and A2. Which of the following represents the most likely change at her β-globin locus and the genetic process that produced it? (Recall that the normal β-globin locus has the gene order Gγ-Aγ- δ-β.)
- Gγ-Aγ-deletion-β, gene conversion
- Gγ-Aγ-β-deletion, gene conversion
- Gγ-Aγ-fusion-δ-β, unequal recombination
- Gγ-Aγ-δ-β fusion, unequal recombination
- Gγ-Aγ-δ-β-Gγ-Aγ-δ-β, equal recombination
7. A 25-year-old male presents for evaluation of declining athletic performance and Physical examination shows mild tachycardia and a palpable spleen, followed by blood counts revealing low hemoglobin (11.7 g/dL) and rounded red blood cells with dense centers on blood smear. A diagnosis of spherocytosis (MIM*182900) is made, and DNA analysis for mutations in the causative ankyrin gene performed, so prenatal or early neonatal diagnosis would be possible for future generations. This mutation is best detected by which of the following?
- Isolation of DNA from red blood cells followed by polymerase chain reaction (PCR) amplification and restriction enzyme digestion
- Isolation of DNA from blood leukocytes followed by Southern blot analysis to detect ankyrin gene exon sizes
- Isolation of DNA from blood leukocytes followed by DNA sequencing of ankyrin gene introns
- Isolation of DNA from blood leukocytes followed by complete gene sequencing or polymerase chain reaction (PCR) amplification/allele-specific oligonucleotide (ASO) hybridization
- Western blot analysis of red blood cell extracts
Questions 8 to 15
Refer to the figure below for these questions.
8. A 3-year-old boy immigrant from Somalia presents for evaluation of priapism (prolonged and painful erection). The parents report through a translator that he has an illness well known in their family, and that several cousins are affected. The illness causes fatigue, poor growth, and bone pain, and the physician suspects sickle cell anemia (MIM*603903). Sickle red cells are observed on peripheral smear, and the physician suggests confirmation by DNA diagnosis so the couple can consider preimplantation/prenatal diagnosis with future.
The β-globin gene is diagrammed in the upper portion (A) of the figure, consisting of untranslated RNA regions (gray boxes), three exons (clear boxes), and two introns (lines between exons). DNA diagnosis for sickle cell anemia was facilitated by finding that a cleavage site for restriction endonuclease (MstII) included the sixth codon of β-globin gene exon 1 (solid circle in first exon). When this codon is mutated to cause the glutamate to valine amino acid change in sickle cell anemia, the MstII site is ablated. MstII sites in normal β-globin genes are shown as solid arrows in the upper part of the figure, yielding gene fragments of 165 bp (5′ MstII site to that in the glutamate codon) and 515 bp (MstII site in glutamate codon to the 3′ MstII site). Recall that the β-globin gene is on chromosome 16, and that both copies are mutated (homozygous) in individuals with sickle cell anemia. Analysis of β-globin gene structure and expression in an individual with sickle cell anemia would yield which of the following results? (Southern blotting is performed using MstII endonuclease cleavage and hybridization with the β-globin gene probe shown in the figure.)
- M stII DNA cleavage segment of 515 and 165 bp by Southern blot, nucleic acid segment of ∼550 bp by Northern blot, and abnormal protein by hemoglobin electrophoresis
- M stII DNA cleavage segments of 515 and 165 bp by Southern blot, no nucleic acid segment detected by Northern blot, and abnormal protein by hemoglobin electrophoresis
- M stII DNA cleavage segments of 515 and 165 bp by Southern blot, nucleic acid segment of ∼1400 bp by Northern blot, and abnormal protein by hemoglobin electrophoresis
- M stII DNA cleavage segment of 680 bp by Southern blot, nucleic acid segment of ∼550 bp by Northern blot, and abnormal protein by hemoglobin electrophoresis
- M stII DNA cleavage segments of 680, 515, and 165 bp by Southern blot, nucleic acid segment of ∼550 bp by Northern blot, and abnormal protein by hemoglobin electrophoresis
9. An 18-month-old Caucasian boy of Italian ancestry is evaluated for “failure to thrive”—his height and weight have leveled off and have gone from the 50th to the 3rd percentiles for Parental neglect is suspected and he is hospitalized for calorie counts and daily weights to see if normal feeding results in weight gain. Upon physical admission, a medical student notes he is pale with brownish-yellow pigmentation to his skin, and that his face seems unusual. Because of these observations, laboratory testing is initiated before the dietary treatment is completed and reveals a low hemoglobin (5 g/dL with normals of 12-14 for age) and target cells on smear that suggest a diagnosis of β-thalassemia (MIM*141900). Molecular studies show a mutation just upstream (5′) to the transcription initiation site of both β-globin alleles (point a in the figure above Questions 8 to 15). In homozygous individuals, this mutation decreased the amount of β-globin mRNA and subsequent β-globin protein and hemoglobin, producing anemia. Which of the following best describes this mutation?
- It affects a promoter sequence that codes for RNA polymerase, lowering β-globin mRNA levels.
- It affects a promoter sequence and the rate at which RNA polymerase II initiates transcription, lowering β-globin mRNA levels.
- It affects the termination site of an upstream gene, increasing β-globin mRNA levels.
- It alters a sequence encoding a subunit of RNA polymerase II (the sigma factor), increasing α-globin mRNA levels.
- It affects a promoter sequence and the rate at which RNA polymerase II terminates transcription, increasing α-globin mRNA levels.
10. A 24-year-old African American mountain climber in excellent physical condition suffers shortness of breath and low oxygen (hypoxia) at high altitude in After transport to base camp and oxygen treatment, a family history reveals that his mother has sickle cell anemia (MIM*603903). With reference to the upper portion (A) of the figure above Questions 8 to 15, laboratory studies of his β-globin gene structure and expression would be expected to show which of the following results? (Note that the same MstII restriction and β-globin probe in the figure is used for Southern blotting.)
- MstII DNA cleavage segment of 515 and 165 bp by Southern blot, RNA segment of ∼550 and 700 bp by Northern blot, and normal and abnormal proteins by hemoglobin electrophoresis
- MstII DNA cleavage segments of 680, 515, and 165 bp by Southern blot, RNA segment of ∼550 and 700 bp by Northern blot, and normal and abnormal proteins by hemoglobin electrophoresis
- MstII DNA cleavage segments of 515 and 165 bp by Southern blot, RNA segment of ∼1400 bp by Northern blot, and single abnormal protein by hemoglobin electrophoresis
- MstII DNA cleavage segment of 680 bp by Southern blot, RNA segment of ∼700 bp by Northern blot, single normal protein band by hemoglobin electrophoresis
- MstII DNA cleavage segments of 680, 515, and 165 bp by Southern blot, RNA segment of ∼1400 and 700 bp by Northern blot, and single abnormal protein by hemoglobin electrophoresis
11. An 8-year-old African American girl presents with severe anemia, despite several years of treatment with iron supplementation. A blood smear is examined carefully and reveals both target cells suggestive of β-thalassemia (MIM*141900) and sickle cells suggestive of sickle cell anemia (MIM*603903). Referring again to the figure above Questions 8 to 15, molecular analysis demonstrates a mutation at the promoter site (point a in the figure) on one β-globin gene and a sickle cell mutation on the Which of the following laboratory results would be expected in such an individual, using the same MstII restriction and β-globin probe for Southern blotting described previously?
- MstII DNA cleavage segment of 515 and 165 bp by Southern blot, normal amounts of ∼700 bp RNA segment by Northern blot, and normal and abnormal proteins by hemoglobin electrophoresis
- MstII DNA cleavage segments of 680, 515, and 165 bp by Southern blot, decreased amounts of ∼550 and 700 bp RNA segment by Northern blot, and normal and abnormal proteins by hemoglobin electrophoresis
- MstII DNA cleavage segments of 680, 515, and 165 bp by Southern blot, decreased amounts of a ∼550 and 700 bp RNA segment by Northern blot, and mostly abnormal proteins by hemoglobin electrophoresis
- MstII DNA cleavage segment of 680 bp by Southern blot, normal amounts of a ∼550 and 700 bp RNA segment by Northern blot, and mostly normal proteins by hemoglobin electrophoresis
- MstII DNA cleavage segments of 680, 515, and 165 bp by Southern blot, lower amounts of 550 and 700 bp DNA segment by Northern blot, and mostly normal proteins by hemoglobin electrophoresis
12. A population study in a rural area of Greece examined several patients with anemia and revealed a homozygous mutation in a sequence 5′-TATAAAA-3′ at the 5′ end of the β-globin gene (site a in the figure above Questions 8 to 15. This sequence has been found at the 5′ boundary of other eukaryotic genes, and is quite similar to a consensus sequence observed in prokaryotes. Which of the following best describes the significance of this mutation?
- Likely β-thalassemia due to disruption of RNA polymerase III binding
- Likely hemoglobinopathy such as sickle cell anemia due to promoter disruption
- Likely β-thalassemia due to disruption of transcription termination
- Major binding site of RNA polymerase I
- Likely β-thalassemia due to disruption of transcription initiation by RNA polymerase II
13. A 2-year-old Caucasian from a Greek community in Chicago presents with severe anemia, targeted red blood cells on peripheral smear, skin pallor, jaundice, and growth Molecular analysis demonstrates a homozygous mutation in the β-globin gene at the junction of exon 1 and intron 1 (site c in the figure above Questions 241 to 248). Which of the following best describes the nature and clinical consequence of this mutation?
- Altered splice donor site, absent mRNA by Northern blotting, and β-globin protein deficiency presenting as α-thalassemia
- Altered splice acceptor site, altered mRNA size by Northern blotting, and β-globin protein deficiency presenting as α-thalassemia
- Altered splice donor site, altered mRNA size by Northern blotting, and β-globin protein deficiency presenting as β-thalassemia
- Altered promoter site, altered mRNA size by Northern blotting, and β-globin protein deficiency presenting as β-thalassemia
- Altered promoter site, deficient β-globin mRNA by Northern blotting, β-globin protein deficiency presenting as α-thalassemia
14. A 22-year-old male is found to have a hemoglobin of 11.5 during his army induction physical and his red cell indices include a mean corpuscular volume (MCV) of 79 (mean 90 femtoliters, 80 is –2 SD). The microcytosis and anemia first suggest iron deficiency, but ferritin levels (reflection of total body iron) are A diagnosis of hemoglobin H disease (MIM*141800) is considered, recalling that the α-globin genes are similar in structure to the β-globin genes diagrammed in the figure above Questions 8 to 15, but are present in duplicate copies on each chromosome 16 in humans. Which of the following α-globin mutations would be compatible with moderate anemia of the type seen in hemoglobin H disease α-globin genes?
- Heterozygous point mutation within the TATA box of one α-globin gene
- Homozygous point mutation within the TATA box of one α-globin gene
- Homozygous mutation deleting both α-globin gene copies
- Heterozygous mutation deleting both α-globin gene copies paired with a heterozygous mutation with the TATA box of one α-globin gene
- Homozygous frameshift mutation within the coding sequence of the 5′ α-globin gene
15. A population survey of a Northern Italian population reveals a variety of mutations in the β-globin It is known that eucaryotic mRNAs undergo several forms of posttranscriptional processing, and that some forms of thalassemia are due to incorrect processing of the α- and β-globin mRNAs. Referring to the β-globin gene structure shown in the figure above Questions 8 to 15, which of the following homozygous mutations would most likely present as altered hemoglobin (hemoglobinopathy) but not as a β-thalassemia?
- Mutations changing the consensus AGGUAAGU splice donor sequence at exonintron junctions
- Missense mutations in exon 2
- Mutations changing the AAUAA recognition sequence at the terminus (3′-end) of the gene
- Mutations altering TATA or CAAT boxes
- Mutations changing the consensus UACUAAC-30bp-CAGG splice acceptor sequence at intron-exon junctions
16. An 18-year-old Ashkenazi Jewish female had developed worsening balance and coordination (ataxia) with declining vision at She also had chronic diarrhea that had been attributed to irritable bowel syndrome but was recognized as fat malabsorption from gastroenterology studies. A diagnosis of autosomal recessive abetalipoproteinemia (MIM*200100) was considered and analysis of apolipoprotein B (apoB) gene expression was initiated. Northern blots of the patient’s duodenal biopsy tissue showed an apoB mRNA of expected size, while Western blots demonstrated an apoB peptide that was smaller than that characterized in control liver tissue. A nonsense, chain-terminating mutation was thought likely until the same small peptide was found in intestinal biopsies of controls. Which of the following interpretations is most likely?
- ApoB mRNA undergoes alternative splicing in intestine; the patient has a heterozygous apoB gene deletion.
- An exon of the apoB gene is deleted in intestine; the patient has homozygous apoB gene deletion.
- Intestinal apoB mRNA undergoes a codon alteration that causes termination of translation and a smaller peptide; the patient may have homozygous apoB missense mutation.
- Polyadenylation of apoB mRNA is deficient in intestine; the patient has heterozygous apoB mutation affecting transcription termination.
- Transcription of apoB mRNA occurs from a different promoter in intestine; the patient has heterozygous apoB mutation deleting this promoter.
17. A 21-year-old Hispanic female presents for evaluation of recurring kidney stones, leg pain, and blanching of her hands with the cold (Raynaud phenomenon). Her prior care was in Mexico and records of prior stone episodes (urolithiasis) or the nature of the stones are not Laboratory testing shows elevated blood urea nitrogen (BUN) and creatinine suggesting chronic renal disease, and urinary excretion of oxalic acid is greatly elevated. A diagnosis of primary oxalosis or hyperoxaluria (MIM*259900) is suggested, known to be caused by abnormal location of alanine-glyoxylate aminotransferase enzyme in the endoplasmic reticulum (ER) rather than the peroxisome. Patients can be treated by liver transplant to restore a source of peroxisomal enzyme, but a less invasive cure could be achieved by engineering one mutant allele so its product enzyme was not targeted to the ER. This alteration would be achieved by which of the following strategies?
- Cleaving the enzyme’s carboxy-terminal segment
- Changing RNA splicing to include an extra exon in the mRNA
- Adding a proteolytic cleavage site near the protein terminus
- Changing the enzyme’s protein processing to produce a smaller peptide
- Altering the enzyme’s amino-terminal sequence
18. A 72-year-old Caucasian male experiences progressive shortness of breath, and is diagnosed with emphysema. He had smoked up to one pack per day of cigarettes until 5 years Family history reveals that his father, two paternal uncles, and his father’s mother all died of emphysema, and a diagnosis of α1- antitrypsin deficiency (AAT-MIM*107400) is suspected. He is found to have decreased amounts and abnormal mobility of AAT protein in his serum when analyzed by serum protein electrophoresis. Liver biopsy discloses mild scarring (cirrhosis) and demonstrates microscopic inclusions due to an engorged endoplasmic reticulum. Which of the following is the most likely explanation for these findings?
- Defective transport from hepatic ER to the plasma
- A mutation affecting the N-terminal methionine and blocking initiation of protein synthesis
- A mutation affecting the signal sequence
- Defective structure of the signal recognition particles
- Defective energy metabolism causing deficiency of GTP
19. A 6-month-old Caucasian boy has had chronic yeast diaper rash and four episodes of pneumonia, two with positive blood cultures for Group A streptococcus. Knowing that Group A strep infections are rare in infants and that the patient likely has defective B-cells that fight bacteria and defective T-cells that fight viruses and fungi, his physician thinks a diagnosis of severe combined immune deficiency (SCID) due to adenosine deaminase (ADA) deficiency (MIM*102700) is Which of the following treatment options would permanently cure the patient?
- Germ-line gene therapy to replace one ADA gene copy
- Germ-line gene therapy to replace both ADA gene copies
- Somatic cell gene therapy to replace one ADA gene copy in circulating lymphocytes
- Somatic cell gene therapy to replace both ADA gene copies in circulating lymphocytes
- Somatic cell gene therapy to replace one ADA gene copy in bone marrow lymphoblasts
20. A 5-year-old Egyptian boy receives a sulfonamide antibiotic as prophylaxis for recurrent urinary tract infections. Although he was previously healthy and well nourished, he becomes progressively ill and presents to your office with pallor and irritability. A blood count shows that he is severely anemic with jaundice due to hemolysis of red blood Which of the following is the simplest test for diagnosis?
- Northern blotting of red blood cell mRNA
- Enzyme assay of red blood cell hemolysate
- Western blotting of red blood cell hemolysates
- Amplification of red blood cell DNA and hybridization with allele-specific oligonucleotides (PCR-ASOs)
- Southern blot analysis for gene deletions