Mammalian Cytogenetics Chromosome structure & karyotype Gene 405: Mammalian Genetics Fall September, 2002

What is cytogenetics? Cyto: Genetics: Study of ………… component of the cell? And that component named as:

What sort of answers does/can it provide? Your feed-back!

Diploid numbers of some commonly studied organisms (as well as a few extreme examples) Homo sapiens (human)46 Mus musculus (house mouse)40 Zea mays(corn or maize)20 Drosophila melanogaster (fruit fly)8 Xenopus laevis (South African clawed frog)36 Caenorhabditis elegans (microscopic roundworm)12 Equisetum arvense (field horsetail, a plant)216 Saccharomyces cerevisiae (budding yeast)32 Canis familiaris (domestic dog)78 Arabidopsis thaliana (plant in the mustard family)10 Myrmecia pilosula (an ant)2 Parascaris equorum var. univalens (parasitic roundworm) 2 Cambarus clarkii (a crayfish)200

Diploid numbers of some commonly studied animals Common Name Genus and Species Diploid Chromosome Number BuffaloBison bison60 CatFelis catus38 Cattle Bos taurus, B. indicus 60 Dog Canis familiaris 78 DonkeyE. asinus62 GoatCapra hircus60 Horse Equus caballus 64 HumanHomo sapiens46 PigSus scrofa38 SheepOvis aries54

Recorded maximum number of chromosomes: Ophioglossum reticulatum This fern has roughly 630 pairs of chromosomes or 1260 chromosomes per cell. Polyploidy is a common in these plants Remarkable!! The cells can accurately segregate these enormous numbers of chromosomes during mitosis.

Recorded minimum number of chromosomes : ant subspecies Myrmecia pilosula Females have a single pair of chromosomes. Males of this group of ants have a single chromosome Reproduces by a process called haplodiploidy fertilized eggs (diploid) become females unfertilized eggs (haploid) develop into males.

Chromosome Three clues to tell chromosomes apart: 1. Chromosome size 2. Position of the centromeres (see also below) 3. Characteristic banding patterns (caused by staining the chromosomes with dyes)

chromosomes are too elongated and tenuous MOSTLY each chromosome is duplicated Before cell division duplicated chromosomes are called dyads Duplicates are held together at the centromere. When first seen contains over 3 million base pairs of DNA. Most of this is repetitive DNA: short sequences (e.g., 171 bp) tandemly repeated Human centromeres complex of proteins that forms at the centromere (11 in budding yeast) Kinetochore sister chromatids, Duplicated chromosomes Shorter of the twop-arm Longerq-arm Arms

The centromere is visible as the primary constriction of the chromosome necessary for correct segregation during cell division site of spindle attachment to the kinetochore and the site of sister chromatid association Primary constriction

Metacentric Sub-metacentric acrocentric

Secondary constriction Nucleolar organizer region (NOR) A chromosomal segment containing genes that encode ribosomal RNA Located at the secondary constriction of some chromosomes Chromatin not visible at these sites NOR bearing chromosomes often bear ‘satellite region’; SATELLITE CHROMOSOMES Length dependent on stage of cell cycle Typically visible from prometaphas through interphase Common cause of erroneous counting of chromosomes

Cellular DNA Nucleus DNA Histones Non-histones Low mol. weight proteins Each chromosome: Cytoplasm How to get the best estimate of the size of haploid nuclear genome? CHROMATIN - avidly staining nuclear body 2/3 1/3 High binding affinity to DNA H1 H2A H2B H3 H4 Structure evolutionarily Highly conserved among eukaryotes High mobility group proteins Insoluble scaffold proteins 5% of total chromosome mass Chromosome core

linker H1 9 fold compaction 5 fold compaction

Numerical description of chromosome shape Chromosome parameters Relative length Centromeric index Arm ratio Descriptors used to group/classify chromosomes Useful in karyotype development Each chromosome: Characteristic length Typical centromere position Conserved size of short (p: petite) & long arm (q)

All chromosomes in a metaphase spread are not of the same size Sizing chromosome in relation to each other Measuring length of entire compliment Finding fraction/percentage for each Relative length (RL) = 100 x length of a chromosome length of haploid compliment for that spread Estimated for several spreads Species standard worked out (used as guidelines)

Chromosome# Genes# of Bases Chromosome million bases Chromosome million bases Chromosome million bases Chromosome million bases Chromosome million bases Chromosome million bases Chromosome million bases Chromosome million bases Chromosome million bases Chromosome million bases Chromosome million bases Chromosome million bases Chromosome million bases Chromosome million bases Chromosome million bases Chromosome million bases Chromosome million bases Chromosome million bases Chromosome million bases Chromosome million bases Chromosome million bases Chromosome million bases Chromosome X million bases Chromosome Y23151 million bases

CENTROMERE INDEX Indicative of where the centromere is located Metacentric chromosomes approx. 50 Sub-metacentric chromosomes < 50 Centromeric index (CI) = 100 x length of short arm total chromosome length Arm ratio (AR) = length of long arm length of short arm If both arms equal: AR is 1 If submetacentric: AR is > 1

Karyotype The complete set of chromosomes in the cells of an organism is its karyotype. Karyotype of the human female Karyotype of the human male 22 pairs of autosomes 1 pair of X chromosomes the same 22 pairs of autosomes one X chromosome one Y chromosome

From banding to schematic representation to idiogram

Chromosome number arm region band 1p32 1p32 1q12 1q31-42

Human idiogram Idiogram Album: Human copyright © 1994 David Adler Resolution

Different types of banding

R-banded metaphase spread

C-banding

Q-banding

M-FISH METAPHASE SPREAD AND KARYOTYPE

How to define a karyotype? Arrangement of all the chromosomes of a cell - based on size, shape and centromeric position - homologous chromosomes arranged together - according to an internationally agreed system of arrangement for the species - accurate identification of homologues (banding) - following international nomenclature Traditional definition Refined definition:

Types of chromosomal aberrations Numerical Structural Euploids Aneuploids

Human trisomies Trisomies involving:211318X161? What do these trisomies share with each other? A number of abnormal characteristics Longterm survival low survival rate survive Fertility a problem ? Mental retardation Retarded growth Cardiac defects Defects/deformities involving different organs/tissues Physical deformities

Relatively frequent Humans & other mammals Structural aberrations Somatic line Where can chromosomes break? When can chromosomes break? What if breakage is in Germ line Deletions Duplications Translocations Inversions

Structural aberrations Deletion Duplication Inversion

Reciprocal Translocation

Clinical significance of chromosome aberrations - Early embryonic death - Viability ? - Spontaneous abortions - Prenatal death - Post natal death (low survivability) - Congenital defects or deformities - Reduced life expectancy - fertility - Abnormalities in offspring abnormal gamete formation

Polytene Chromosomes Giant chromosomes - dipteran Begin as normal chromosomes Repeated rounds of DNA replication (endoreplication); & no cell division Become large banded chromosomes Centromere – no endoreplication Bundle in a mass - chromocenter Found in larvae - allow faster growth Bands unique to each chromosome Allow high resolution mapping