Transcript ppt presentation

Chromosomes and
chromatine
structural arrangement of genetic
information
Chromatin (Walther Flemming 1882)
= DNA + associated proteins
euchromatin x
heterochromatin
heterochromatin facultative x constitutive
Chromatin
(DNA + associated proteins)
Genetic information = DNA sequence (change = mutation)
- protein-coding, regulatory, RNA-coding
-
Epigenetic information (less stable, depends on location)
- transcriptional activity, access of interacting proteins
transcriptionally active
decondensed, accessible
x
x
transcriptionally inactive
compact, unaccessible
Epigenetic modifications of chromatin
- epigenetic information can be mitotically and meiotically
herritable (e.g. some changes in gene activity)
- no change in primary DNA sequence
- modifications of chromatin components:
• DNA methylation
• histone posttranslational modifications mutually interconnected!
• histone types
euchromatin
heterochromatin
Nucleosome
octamer of histones (small alcaline proteins):
2 x H2A, 2 x H2B, (2 x H3, 2 x H4)
+ 147 bp DNA
Varying composition:
- histone variants (isoforms): CenH3, H3.3, H2A.Z
- posttranslational modifications of histone proteins
Structure of 30nm fibre
Solenoid or ZigZag?
- still unclear
solenoid
Li and Reinberg 2011
Higher structural order of chromatine
- hypothetically loops with actively transcribed genes
- insufficiently understood
• MARs (matrix attachment regions)
alt. SARs (scaffold attachment reg.)
– harboring regions surrounding coding
sequences to nuclear protein matrix
– AT rich, colocalize with „insulators“
(sequences that prevent spreading of
heterochromatin)
– distances between 3 - 100kb
• LBARs
(loop basement attachment regions)
- organize chromosomes to huge loops
(distances 20kb až 100kb)
example of hypothetical
arrangement
Interconnections betweeen
nucleosomes
- linker sequence between: length = multiple of 10 bp
(20 to 90 bp)
- average (most frequent) length - differences among
species, tissues, …
(20 bp yeast, 30 bp Arabidopsis, 40 bp mammels)
- internucleosomal fragmentation yields: 167 – 237 bp
( frequent length of repeats)
Interconnections betweeen nucleosomes
Direct interactions
- N-ends of H4 interact with H2A.H2B bodies in parallel fiber
- presence of H2A.Z variant probably prevents parallel
interaction
Linker histone H1
- alcaline both ends (amino and carboxy)
interaction with both histones and DNA
- stabilization of higher structures (30nm), phosphorylated
during cell cycle
- length of linker sequence:
longer - require H1, more compact –
heterochromatin
shorter – H1 less important, more decondensed,
active chromatin
Nucleosome position
• arrangement on DNA is not random (but is changable)
- DNA sequence
- DNA methylation
- histone modification/types
- DNA transcription
• regulation / modulation of transcription
- „unstable nucleosom region“ (earlier „nucleosom-free region“) in front
of transcription start site (mainly constitutively expressed genes) –
- unstable nucleosomes with H3.3 and H2A.Z histones
- surrounded with stably situated nucleosomes with H2A.Z
- nukleosomes help to define exons (central location even without
transcription!)
Histone code
- covalent posttranslatinal modifications (PTM)
- modifications mainly on N-ends (out of core)
- high complexity
- „epigenetic instruction“ to manage with DNA
Some histone PTMs are mutually
interconnected and have multiple functions
e.g. H2A phosphorylation – injured DNA labelling,
but also role in regulation of transcription and spiralization
and in PCD
Rossetto et al. 2012
Histone code
Phosphorylation – predominantly short-time transient label, various
functions
Acethylation – predominantly „executive modification“ for weakening
interaction with DNA
- K-Ac – specific interaction of bromodomain proteins
– signal role ( stabile), both repressive and activating (~
depends on position)
- K-Me – specific interaction with chromodomain and TUDORlike domain proteins
- key role in regulation of DNA methylation
and chromatine activity
- H3K9me2, H3K4me3, H3K27me3
Methylation
Reproduction of nucleosomes after
replication
- histone tetramers (H3/H4) and dimers (H2A.H2B) not
divided between sister strands!
- one strand – „parental histons“ (Asf1) de novo deposition
(CAF1, Asf1)
- H2A.H2B incorporated even later (post replication)
Chromosomes
NOR: 18S- 5,8S- 26S rDNA
Caryotype – number, types and sizes of chromosomes
Classical caryotype
(metaphase)
Flow caryotype
(FISH labelling)
Doležel et al. 1999
Chromosome number and sizes
Number: 2 - 600
Size: 2,4 Mb Genlisea
30 Mb Arabidopsis
800 Mb Triticum
What are the consequences?
- different linkage groups
(various gene recombination)
- limited hybrid fertility, …
Chromosome number differs
between species
• Extreme chromosome numbers
– Haplopappus gracilis: 2n = 4
– Sedum suaveolens: 2n = cca 640
• Luzula sp.:
– 2n = 6 až 66
– holocentric chromosomes
– Chromosom size differes up to 60x
L. pilosa
(Cullis, Plant genomics and proteomics, 2004)
L. elegans
B chromosomes in plants
- non-pair chromosomes in some species (1500 species – maize)
- usually no protein-coding genes
- usually negatively affect fitness (fertility)
- not present in all individuals in population
- parazitic DNA (?)
Chromosome number and genome size
Telomeres
DNA-protein structures serving to
maintain stability of chromosomal
ends
Repetitive sequences synthetized by
telomerase after replication
(TTTAGGG)n in Arabidopsis
Some plants have typical mammalian
sequence: (TTAGGG)n
Keeping telomere length
Telomerase - RT with an RNA template
repeat number depends on:
- species
- developmental stage
- cell type
- chromosome (within a cell)
Centromeres
- attachement of chromatids
- defined by presence of histone CenH3
- CenH3 – kinetochore – spindel fibers
Types:- point (125 bp, yeast Saccharomyces)
- holocentric chromosomes (CenH3 along whole chr.)
e.g. Luzula – allows fragmentation
- classical – region of different length formed with
heterochromatin (repeats, TE)
= epigenetically defined (neocentromeres)
- various strenth in hybrids
Chromosomes: ((telocentric, acrocentric, metacentric,submetacentric))
Crossing of WT Arabidopsis with a line carrying modified
(weaker) CenH3 issues in haploid progeny
– inefficient segregation of chromosomes (elimination)
Ravi and Chan 2010
Chromosomal territories
Regions in nucleus occupied with certain chromosome
(postmitotic decondensation 2 hours, 2.5 fold increse)
Rabl’s arrangement of
chromosomes in
interphase nuclei
centromeres and
telomeres localized in
oposite sites
(chrom. size above 500 Mb)
WHY?
Experimental confirmation of chromosomal territories
- specific labelling
of chromosomes (FISH)
- laser injury, detection of reparation
Cremer and Cremer 2010
Sex chromosomes in plants
- sex determination (single locus or more loci)
Sexuality in various taxons of plants evolved independently and
repeatedly (5 % species, in about 75 % plant families)
- Marchantia, Gingo, Silene, Rumex, Hop, Poplar …
Sex chromosomes in plants
Morphological classification of sex chromosomes
- homomorphic
- heteromorphic
- polymorphic – more than two types:
e.g. Rumex acetosa: male XY1Y2, female XX
Humulus lupulus var. cordiflorus: male X1X2Y1Y2, female X1X1X2X2
Evolution of sex chromosomes
Formation of sex chromosome and single sex individuals
– primary mutations causing male and female sterility in loci
in strong genetic linkage (intermediarily usually gynodioecy)
model:
female (XX) – an allele (in locus A) necessary for development of
male sex organs is non-functional in X-chromosome ancestor
(recessive allele)
male XY – an allele (in locus B, linked with locus A) is mutated to
suppress formation of female sex organs (dominant allele), this
allele is linked with functional allele in locus A
Evolutionary young – homomorphic
(recombination only partially limited)
Degenerations (inversions, TE amplification,
deletions) - heteromorphic
Splitting or translocations can issue in
polymorhic
Polyploidy complicates formation of sex chromosomes