Principles of cytogenetics Flashcards
Cytogenetics
study of chromosomes, their structure and inheritance, as applied to the practice of medical genetics.
Chromosome sources
T-lymphocytes, chorionic villi, amniotic fluid. Arrest cells in metaphase since they are condensed and separated
Chromosome nomenclature
chromosome arms (p1, p2, q1,q2 etc) divided into regions counting outward from the centromere. Regions are divided into bands labeled p11, p12 . . . sub-bands: p11.1, p11.2 . . .sub-sub bands p11.21 p11.22 etc
Metacentric
centromere near central region of chromosome
Submetacentric
centromere is off center and arms are clearly of different lengths
Acrocentric*
centromere near one end (13, 14, 15, 21, 22, Y)-must know! Contain small distinctive masses of chromatin called satellites attached to their short arms by narrow stalks which contain hundreds of copies of ribosomal RNA genes. p arm consists only of rRNA; q arm will hold the important genes.
Telocentric
centromere at one end and only has a single arm ( does not occur in a normal human karyotype-unless mutation loss of p arm)
Giemsa banding (G banding)
Most common method;pro/metaphase cells treated with trypsin to digest proteins->Giemsa stain->produces light and dark bands. (400bands)
Quinacrine mustard (Q banding)
stain with fluorescent quinacrine mustard. Examine by fluorescence microscopy. Bright Q bands correspond exactly to dark G bands. Used to detect heteromorphisms/variants in chromosome morphology. reflect diffs in satellite DNA amts
R banding
“Reverse banding” chromosomes receive special treatment such heating before staining. Dark/light band pattern opposite of that observed by G or Q banding. Used to examine regions that stain poorly by G or Q banding. Common in Europe-abns clearly visualized.
T banding
“Telomere” IDs subset of R bands esp concentrated at telomeres. Exteme heat treatment before staining with Giemsa-combo of dyes and fluorochromes.
C banding
Staining CENTROMERIC REGIONS of each chrome and other regions containing CONSTITUTIVE HETEROCHROMATIN
High-Resolution banding (Prometaphase banding)
G or R banding of chromosomes from a prophase or pro metaphase cell. Less condensed chromatin useful to detect subtle structural abns in a chromosome. (850)
Fluorescence In Situ Hybridization (FISH)
CHROMOSOMAL PAINTING-probes for entire chromosome or arm. SPECTRAL KARYOTYPING-can analyze multiple targets simultaneously utilizing diff fluorochromes. Cell culture still req’d, but no need to lyse, just add fluorochromes. Locus specific, satellite DNA, Chromosome paint.
Spectral karyotyping
Use diff colors/fluorochromes for all 24 chromosomes-> can ID abns.
Comparative Genome Hybridization (CGH)
small changes in # of copies of a DNA segment can be identified using CGH, measures the difference b/w 2 diff DNA samples in copy, or dosage of a particular DNA segment.
Microarray analysis
CGH adds info to karyotype, but doesn’t replace. Assess relative copy #, complements karyotyping, more sensitive, analyzes whole genome, but does not give info about structural changes i.e. translocations, rearrangements
Chromosome Abnormalities
Numerical or Structural, may involve autosomes or sex chromes or both.
*Euploidy
chromosome # that is a multiple of n (23)
Haploid is n=23
Diploid is 2n=46; normal somatic cells
*Heteroploidy
any chromosome complement other than 46.
Triploid (3n) and Tetraploid (4n)
euploid and heteroploid at the same time-not compatible with life-seen in abortion.
Triploid
3n, often due to fertilization by 2 sperm -failure of meiotic division -partial hydatidiform moles-> extra paternal -> abnormal placenta extra maternal-> spontaneously aborted
Tetrapoid
4n; failure to complete an early cleavage division of the zygote.(meiosis I)
Aneuploidy
Not a multiple of 23 or haploid #. A result of MEIOIC NONDISJUNCTION.
Monosomy, trisomy
monosomies and trisomies of 6 chromosomes are more compatible with life.
Monosomy
autosomal cell with only 1 copy of a chromosome-lethal
monosomy of sex chromosome-can survive
monosomy of autosome-death of zygote
Trisomy
3 copies of a chromosome- potentially lethal
Nondisjunction
failure of a pair of chromosomes to disjoin properly during one of the 2 meiotic divisions. usually meiosis I.
Nondisjunction during meiosis I
gamete with 24 chromosomes has both maternal and paternal members of pair
Nondisjunction in meiosis II
gamete with 24 chromosomes has either maternal or paternal copies of the pair
Nondisjunction in mitosis
Mosaicism=2 DIFF GENOTYPES DERIVED FROM 1 CELL TYPE->zygote->cleavage divisions->morula->then cells divide and will always have the nondisjunction. So some cells will have trisomies and some will have normal.
Abnormalities of chromosome structure: result from . . .
result from chromosome breakage followed by reconstitution in an abnormal combination.
Less common, spontaneous, induced by BREAKING AGENTS (clastogens, viral infections, ionizing rad, chems)
may be in all cells or mosaic
balanced-> chrome set has normal complement of chromosomal material
unbalanced-> additional or missing material
Abnormalities of chromosome structure: stablel rearrangements
Stable: capable of passing unaltered through mitotic and meiotic divisions. STABILITY REQS NORMAL STRUCTURAL ELEMENTS-including a fxnl CENTROMERE and 2 TELOMERES.
Abnormalities of chromosome structure: Unbalanced rearrangements
Phenotype likely to be abn due to deletions, duplications, or both. Important class->submicroscopic changes involving telomeres.
Idiopathic Mental retardation: may be diagnosed by targeted cytogenetic analysis of telomeric regions by FISH
Unbalanced rearrangements:deletions
clinical consequences due to haploinsufficiency.
TERMINAL or INTERSTITIAL
due to chromosomal breakage and loss of acentric segment, unequal crossing over, abn segregation of a balanced translocation or inversion. detected by FISH or high-resolution banding
Unbalanced rearrangements:duplications
results in partial trisomy
due to unequal crossing over, ban segregation of a translocation or inversion.
Unbalanced rearrangements: marker and ring chromosomes
small unidentified chromosomes often seen in mosaicism. supernumerary chromosomes or extra structurally abn chromes. Dx with FISH. Commonly seen with chrome 15 and sex chromes.
Unbalanced rearrangements: marker and ring chromosomes. NEOCENTROMERE
Neocentromere: subclass of marker chromosomes; small fragments of chromosomes that do not have centromeric seqs; mitotically stable, has acquired centromere activity
Unbalanced rearrangements: Ring Chromosomes
marker chromes without telomeres; formed by loss of terminal ends of both arms and the ends join together; rare-mitotically stable.
Unbalanced rearrangements: Isochromosomes
chromes in which one arm is missing and the other duplicated in a mirror image fashion.
misdivision through centromere in meiosis II; exchange involving one arm of a chrome and its homolog at the proximal edge of the arm adjacent to centromere. MOST COMMONLY OBSERVED: ISOCHROMOSOME X(involving q arm)
Unbalanced rearrangements: Dicentric chromosomes
Rare: 2 chrome segs each with a centromere fused end to end.
Psuedocentric: can be mitotically stable if the 2 centromeres are coordinated in their movement during anaphase or one centromere is inactivated.
Balanced rearrangements
usually no phenotypic effect in carriers; likely to produce unbalanced gametes -> increased risk of abn offspring.
Ex: X-linked diseases in female carriers of balanced X: autosome translocations
Balanced rearrangements: Inversions
single chromosome undergoes 2 dbl strand breaks and is reconstituted with the segment b/w the breaks inverted.
Balanced rearrangements: Inversions-Paracentric
Not involving centromere-> unbalanced gametes with a centric or dicentric recombinant chromes not viable-> risk of live born with karyotype is low
Balanced rearrangements: Inversions-Pericentric
Involves centromere->unbalanced gametes with duplication and deficiency of segments distal to inversion. Increased risk of abn offspring
Pericentric inversions of Chormosome 3
Carriers of inv(3) are normal, but offspring have characteristic abn phenotype.
Pericentric inversion of chromosome 8
carriers of inv(8) have 6% chance of child with recombinant 8 syndrome->lethal disorder with severe cardiac abns and mental retardation.
Most common Pericentric inversion
inv(9)(p11q12) . . . considered a normal variant
Balanced rearrangements: Translocations
Most important type! Exchange of chromosome segments b/w non-homologous chromosomes (i.e.9 and 10)
Reciprocal Translocation (balanced)
break in 2 diff chromes and material exchanged->unbalanced gametes and abn progeny.
Robertsonian Translocation
Long arms (q arm) of 2 homologous *acrocentric chromes- fuse at the centromere forming a single chromosome and p arm is lost. Balanced karyotype with 45 chromes. Clinically->carriers for chromosome 21 have risk of producing child with Down syndrome-young mothers
Balanced rearrangements: Insertions
nonreciprocal, segment removed from one chrome is inserted into a diff chorme. Reqs 3 breaks. 50% risk of abn offspring.
Mosaicism
2 or more chromosome complements in an individual-> structural-> commonly nondisjunction in a post zygotic mitotic division.
Abns can be seen in all tissues, but only some of the cells exhibit the abnormal phenotype.
Pseudomosaicism
mosaicism that arose in culture
Spontaneous abortions
20% of cases 45,X . . . overall rate is 15%
Genomic imprinting (unusual inheritance patterns)
diffs in gene expression b/w the allele inherited from the mother and the allele inherited from the father. Caused by changes in chromatin like methylation or cytosines, modification or substitution of specific histone types-> affects the expression of the gene, but not the DNA seq.
Characteristic feature: only 1 allele is expressed.
Epigenetic mechanism
factors that affect gene fxn without changing the genotype; methylation, histone modifications, chromatin structue etc.
Affect gene expression without changing primary seq.
Genomic imprinting: Prader-Willi and Angelman syndromes
Genomic imprinting diseases involving the SAME CHORMOSOMAL LOCATION:15q11q13.
Key diff: whether the abn chrome is maternally/paternally derived!!
If ABN CHROMOSOME 15 is inherited from:
Father-Prader-Willi; patients’ genetic info derived only from their mothers
Mother-Angelman; patients’ have genetic info derived only from their fathers
Usually 2 cytogenetically normal chromes both inherited from one parent
Prader-Willi syndrome phenotype
Obesity, excessive and indiscriminate eating habits, small hands and feet, short stature, hypogonadism, and mental retardation
Angelman syndrome phenotype
Unusual facial appearance, short stature, severe mental retardation, spasticity, and seizures
Uniparental Disomy
Minority of PWS and AS cases have inherited both chromosomes from one parent:
PWS if both from mother
AS if both from father
Isodisomy
if identical chromosome is present in duplicate
Heterodisomy
if both homologs from one parent present
Uniparental Disomy: Beckwith-Wiedemann syndrome
Infants large at birth, enlarged tongue, frequent protrusion of umbilicus, severe hypoglycemia, development of kidney, liver, and adrenal malignancies
Causes: excess of paternal or loss of maternal contribution of genes or both on chromosome 11q15, including Insulin-like GF2 gene.
Hydatidiform mole
only paternal contribution; caused by abn growth of chorionic villi-> epithelium proliferates and stoma undergoes cystic cavitation-> placenta converted into a mass of tissues resembling a bunch of grapes.
Occurs when sperm fertilizes an ovum that lacks a nucleus-> duplication of haploid male chromosome set(sperm chromes); no maternal chromes so no fetal tissue at all!
Can lead to choriocarcinoma
Hydatidiform mole patient presents with:
excessive vomitting, height of fundus does not correspond to the date-its higher than expected; enlarged uterus mean abn pregnancy
Hydatidiform mole treatment
Fragile uterus easily ruptured so can’t do endometrial curettage which can cause death. So use suction curettage and slowly suck out the tissue. HCG levels must fall. If still high, not all of mole was removed so must go back and suck it out.
Ovarian teratomas
benign, arise from 46,XX cells containing only maternal chromosomes.
Confined placental mosaicism-placenta has abnormality that is not apparent in fetus, but leads to abnormal fetus.
Mendelian disorders with cytogenetic effects
Chromosome instability syndromes: rare single gene syndromes with characteristic cytogenetic abnormality.
