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Dasar-Dasar Patologi

Subseluler & Molekuler

UPIK A. MISKAD, PhD, SpPA

SEL EUKARIOTIK & PROKARIOTIK

ORGANEL SEL

Inti Sel

Bentuk : bulat atau lonjong


Diameter 3 4 m
Mengandung materi genetik
Tanpa inti sel sel mati
Komponen inti sel :
Selaput inti
Kromatin
Nukleoplasma
Neukleolus

Membran Nukleus

Nukleolus

Kromatin

Membran Plasma
Nama lain : Plasmalema
Tersusun bi-layer
Fosfolipid di bagian tengah dan diliputi
protein
Bagian permukaan hidrofilik
Bagian tengah hidrofobik
Protein membran : protein integral & perifer

Membran Plasma

Mitokondria
Berbentuk filamen selebar 0,5 1 m
Dalam 1 sel bisa terdapat sampai 800
buah (sel hati)
Sumber energi terbesar sel : ADP
ATP
Jumlah krista dalam mitokondria
sesuai aktivitas sel
Terdapat DNA mitokondria

Gambar Mitokondria

Ribosom
Fungsi : merangkai asam amino peptid
Messenger RNA yang keluar dari nuclear
pore diubah menjadi peptida oleh
ribosom (Translasi)
Terdapat dalam sitoplasma dan melekat
pada REK
Terdiri dari 2 sub unit : Besar & kecil

Sintesis Protein

Gambar Ribosom

Retikulum Endoplasma
Kasar
Suatu bangunan berlapis yang
membungkus suatu ruang intersisterna
yang berisi protein-protein yang baru
disintesa
Ribosom menempel pada bagian luar
retikulum endoplasma REK
Fungsi utama : pembentukan protein

Gambar REK

Retikulum Endoplasma
Halus
Tidak terdapat ribosom pada permukaannya
Peran : produksi lipid, metabolisme
karbohidrat, detoksifikasi racun
Sel yang metabolismenya aktif jumlah
REH
Sel hati jumlah REH

Gambar REH

Alat Golgi
Terlihat sebagai jaring-jaring atau vakuol
yang gelap bila dilihat dengan mikroskop
cahaya
3 komponen :
Sisterna
Vesikel kecil
Vakuol besar
Di dalam sel hanya terdapat 1 alat golgi
Terdapat receiving site dan shipping
site

Gambar Alat Golgi

Lisosom
Suatu sistem pencerna intrasel yang mampu
memecah materi yang berasal dari dalam dan dari
luar sel
Bentuknya bermacam-macam Pleiomorfisme
Mengandung berbagai macam enzim hidrolase
Terdapat 2 macam : Primer dan Sekunder
Fagosom :Vakuol ok internalisasi bahan dari luar
Autosom : Vakuol internalisasi bahan dari dalam
Autosom / Fagosom + Lisosim primer Lis sekunder

Gambar Lisosom

Peroksisom
Peroksisom atau badan mikro mempunyai
struktur yang sama dengan lisosom, tetapi
tidak mengandung enzim hidrolase
Organel ini dibentuk di dalam REK dan
fungsinya sampai sekarang masih belum
jelas
Diduga fungsi organel ini adalah melindungi
sel terhadap peroksida hidrogen yang
sangat toksik terhadap sel. (mengandung
enzim peroksidase)

Sentriol
Dengan mikroskop cahaya sentriol tampak
sebagai granula atau batang pendek yang
terletak dekat dengan inti
Pada umumnya pada saat interfase sel yang
tidak membelah mempunyai 2 buah sentriol atau
disebut diplosom.
Bila dilihat pada potongan melintang , dinding
silinder tersebut tersusun oleh sembilan triplet
yaitu gabungan 3 mikrotubul yang menyatu.
Triplet tersebut tersusun agak serong terhadap
lingkaran sentriol.
Dan selama mitosis, kedua pasangan yang
dihasilkan bergerak ke kutub berlawanan dari sel
dan menjadi pusat organisasi bagi gelondong
mitosis

Gambar Sentriol

Mikrotubul
Mempunyai garis tengah luar 24 nm,
terdiri atas dinding padat setebal 5 nm
dan lapisan pusat selebar 14 nm.
Subunit sebuah mikrotubul adalah
heterodimer yang terdiri atas molekul
tubulin alpha dan beta.
Peran : gerakan sel , dalam proses
mitosis bila dikaitkan dengan sentriol ,
transpor intrasel

Gambar Mikrotubul

3 Macam
Mikrotubul

Ikatan
Kromosom
dan
Mikrotubul
.

Pembelahan Kromosom atau Anafase

Outline of Gene Expression


Central dogma of molecular biology:

Three main functions of DNA


1.

2.
3.

To store information (genes, regulatory signals, ...)


Gene definition: Physical and functional unit of
heredity which carries information from one
generation to the next.
Molecular biology definition of a gene: It is the entire
DNA sequence necessary for production of a
functional protein or RNA
To replicate faithfully
reliably pass information to daughter cells
Allow for a slow changes over time (mutations)

Overview of DNA structure

How does DNA carry the


information for life?
Small segments of DNA (sequences of
nucleotides) code for specific proteins
These segments of DNA are called
genes
One gene codes for one protein
Hundreds-Thousands of genes per
chromosome

RNA
Three different types of RNA:
mRNA - messenger RNA,
specifies order of amino acids
during protein synthesis
tRNA - transfer RNA, during
translation mRNA information
is interpreted by tRNA
rRNA ribosomal RNA,
combined with proteins aids
tRNA in translation

Messenger RNA (mRNA)


Carries the information for a specific
protein.
Made up of 500 to 1000 nucleotides long.
Made up of codons (sequence of three
bases:
AUG methionine).
Each codon, is specific for an amino acid.

Messenger RNA (mRNA)


start
codon
mRNA

A U G G G C U C C A U C G G C G C A U A A
codon 1

protein methionine

codon 2

codon 3

glycine

serine

codon 4
isoleucine

codon 5

codon 6

glycine

alanine

codon 7
stop
codon

Primary structure of a protein


aa1

aa2

aa3
peptide bonds

aa4

aa5

aa6

Transfer RNA (tRNA)


Made up of 75 to 80 nucleotides long.
Picks up the appropriate amino acid floating in
the cytoplasm (amino acid activating enzyme)
Transports amino acids to the mRNA.
Have anticodons that are complementary to
mRNA codons.
Recognizes the appropriate codons on the
mRNA and bonds to them with H-bonds.

RNA differs from DNA


1. RNA has a sugar ribose
DNA has a sugar deoxyribose
2. RNA contains uracil (U)
DNA has thymine (T)
3. RNA molecule is single-stranded
DNA is double-stranded

DNA & RNA

DNA RNA Protein


Nuclear
membrane

DNA
Transcription

Eukaryotic
Cell

Pre-mRNA

RNA Processing

mRNA
Ribosome
Translation

Protein

Transcription
The transfer of information in the nucleus from a
DNA molecule to an RNA molecule.
Only 1 DNA strand serves as the template
Starts at promoter DNA (TATA box)
Ends at terminator DNA (stop)
When complete, pre-RNA molecule is released.

Three Phases of Transcription in Eukaryotes:


1) Initiation RNA polymerase binds DNA.
2) Elongation - Polymerase reads from the DNA
template strand and builds complementary RNA.
3) Termination - Release of polymerase

Where does transcription initiate?

RNA polymerase
binds to the TATA
box in the promoter,
which is located just
5 (upstream) of the
transcription start
site.

Transcription

Transcription

When the termination site is reached,


the RNA polymerase is released.

RNA Processing
The RNA produced during
transcription is
called pre-mRNA

Pre-mRNA must be processed into


mature
mRNA
This occurs in the nucleus

RNA processing includes:

Addition of a 5 cap (modified GTP)

Addition of a poly-A tail

Splicing to remove introns

Post-transcriptional modifications of RNA

During splicing, introns are cut


out and
exons are spliced together.

Translation
Synthesis of proteins in the cytoplasm
Involves the following:
1. mRNA (codons)
2. tRNA (anticodons)
3. rRNA
4. ribosomes
5. amino acids

aa4

aa5

Termination

aa199

aa3 primary
structure
aa2 of a protein

aa200

aa1
200-tRNA

A C U
mRNA

terminator
or stop
codon

C A U G U U U A G

Codon - Amino Acid

DNA Replication
3

AT
C G
A
T

A .. T
G...C
T ..A

DNA Replication
5
An enzyme termed DNA
Polymerase synthesizes
new DNA from original in a
5 to 3 direction (the end
with a free OH group)
DNA replication results in
two new complementary
strands of DNA.
One of each is from original
and one new..semiconservative replication

3
New nucleotides can
Only add to this end

G
T C
A
A
T
C
G

AT
C G
A
T

DNAPolymerase

5
3

DNA replication involves a great many building


blocks, enzymes and a great deal of ATP energy.
DNA replication in humans occurs at a rate of 50
nucleotides per second and ~500/second in
prokaryotes.

Nucleotides have to be assembled and available in


the nucleus, along with energy to make bonds
between nucleotides. DNA helicase enzymes unzip
the DNA helix by breaking the H-bonds between
bases.

Since the DNA strands are antiparallel, and replication proceeds in the 5' to 3'
direction on each strand, one strand will form a continuous copy. The top strand
here.

The Lagging Strand

while the other, lagging strand will form a series of short pieces
with gaps. These are called Okazaki fragments and require the use
of other enzymes to complete the process.

DNA Damage
DNA Damage is caused by a variety
of agents and by different
mechanisms
Replication errors may cause
mismatched bases
Cells of all types have a number of
DNA repair pathways to deal with
mismatches and damage

DNA Damaging Agents


Irradiation
Chemical
Note: Mutagens change the DNA
sequence but not necessarily by
DNA damage (mostly chemical
change from one base to another
base)

X-ray Irradiation

X rays break the DNA backbone

Oxidation from free radicals


(formed by irradiation)
damages individual bases

Deamination

Depurination

Base Excision
Repair

10 September

Nucleotide Excision
Repair

10 September

The flow genetic information


(Central dogma in Molecular
Biology)
DNA

Replication

DNA

Transcription

RNA
Translation

PROTEIN

REPLICATION (DNA SYNTHESIS)


TRANSCRIPTION (RNA SYNTHESIS)
TRANSLATION (PROTEIN SYNTHESIS)

CELL CYCLES
The
repeating
sequence
of
growth and division through
which cells pass each generation
The purpose of the cell cycle is
to allow cells to reproduce
without alteration of genetic
material
The stages are well preserved
between species

Why Do Cells Divide?

Growth
Development
Maintenance
Reproduction

Cell Cycle Progression


Higly regulated process controlled by a
large number of factors include:
Growth stimulating factors
Growth inhibiting factors
Enzymes that alter functional state of other
proteins by adding (kinases) or removing
(phosphatases) phosphate groups

Failure in the mechanisms by which the


cell cycle is regulated can lead to
uncontrolled cell growth and the
development of cancer

The Cell Cycle Control


There
are
important
during the cell cycle:

checkpoints

at the G1 S transition
at the S G2 transition
at the G2 M transition

Regulation of the cell cycle occurs as a


consequence of cyclin Cdk interaction
Other proteins such as P53, Rb and
oncogene play a role in regulating the
cell cycle

Events of Cell Cycle


G1 Phase Period of growth; for many
organisms this occupies a major portion of the
cells life span
S Phase DNA replicates, resulting in
duplicate chromosome
G2 Phase Cell prepares for division (mitosis),
building necessary materials; centriole and
other organelles double
M Phase Cell division, consisting of nuclear
division and cytoplasmic division (cytokinesis)

Cell Cycle Is Regulated By


Opposing Effects of Positive and
Negative Factors

S
Cyclin

P53
+

Cdk

G2

G1
M

Rb

Cell Cycle Is Regulated By


Opposing Effects of Positive and
Negative Factors
Tumor Suppressor
(P53, Rb)

Oncogene
(BCl2,BRCA1)

Cell death

Cell proliferation
NORMAL

Cell death
Oncogene
(BCl2,BRCA1)

Tumor Suppressor
(P53, Rb)

Cell proliferation
CANCER

Cyclins and Cyclin Dependent


Kinases (Cdk)

Cyclin
Cyclin D
Cyclin E
Cyclin A
Cyclin A
Cyclin B

Cdk
Cdk 4,6
Cdk 2
Cdk 2
Cdk 1(Cdc2)
Cdk 1

Process regulated
G1 phase progression
G1 to S phase
S phase progression
S through G2
M phase

Waves of Cyclin-Cdk Activity


Control
The Cell Cycle
G1

G2

G1

Cyclin B-Cdk1

Kinase Activity

CyclinA-Cdk1
Cyclin E-Cdk 2
Cyclin D-Cdk 4/6

Time
GF

CHECKPOINTS
The cell cycle control system has
checkpoints during G1, G2, S and M
Checkpoint signals report cells
status:

Is the cell big enough?


Is environment favorable?
Is DNA replicated?
Are chromosomes attached to opposite
poles?

G1

G2

prophase

metaphase

anaphase

Mitosis

Cytokinesis

GENETIC VARIATION
AND DISEASE

Concept of Genetic Variation


Genetic Variation or polymorphism
means that a certain gene locus can
have numerous variants or 'alleles
Genetic variations occurring in more
than 1% of a population would be
considered useful polymorphisms for
genetic linkage analysis.
Genetic variation among individuals
means that each of us has a unique
genetic make-up that influences our
susceptibility to disease.

A C T C A G T

T G A

General population

97%

A C T C A G T

T T A

Polymorphism

3%

A C T C A G T

T G A

General population

99.8%

A C T C A G T

T T A
0.2%

Mutation

What is genetic variation?


Different people can
have
a
different
nucleotide or base at a
given location on a
chromosome

. . . . G G T A A C T
G.....
. . . . G G C AAC T G . . . .
. . . . . G G G AAC T G . . .
.

What is an genetic variation map?


Human DNA

Location of genetic
variation on human
DNA

How can the map be used to predict


disease?

Plasma level of ACE are under strong


genetic control and genetic variation in the
ACE gene have been proposed as a risk
factor for coronary heart disease

DD genotype of ACE gene was


associated with coronary heart
disease

ACE
genotype

DD ID

gene

II

THE EFFECTS OF
GENETIC VARIATION
Neutral alteration, a base substitution with
no effect on the amino acid residue
encoded
Conservative change, a base substitution
that results in an altered amino acid, but
has minimal effect on protein structure
and function
Nonconservative alteration leading to a
change in the encoded amino acid residue
that has dramatic effects on protein
structure and function

The Genetic Code


U

UUU
UCU
UUC Phe UCC
U
Ser
UUA Leu UCA
UUG
UCG

UAU
UAC Tyr
UAA Stop
UAG Stop

U
UGU
Cys
C
UGC
UGA Stop A
UGG Trp G

CUU
CCU
CUC
CCC
C
Leu
Pro
CUA
CCA
CUG
CCG

CAU
CAC His
CAA
Gin
CAG

AUU
AUC Ile
A
AUA
AUG Met

ACU
ACC
Thr
ACA
ACG

GUU
GUC
G
Val
GUA
GUG

GCU
GCC
Ala
GCA
GCG

U
AAU
AGU
Ser
C
AAC Asn AGC
AAA
AGA Arg A
Lys
AAG
AGG
G
U
GAU
GGU
C
GAC Asp GGC
Gly
GAA
GGA
A
Glu
GAG
GGG
G

U
CGU
C
CGC
Arg
CGA
A
CGG
G

Positions of mutant sites and


their functional consequences
Wild type

Neutral alteration
Conservative change
Nonconservative change
Nonconservative change

Promotor region
Exons
Mutant site

Components of protein active site


Intron

Functional Consequences of Genetic


Variations
Loss of function
+

Leak of function

Gain of function

LEVEL OF VARIATION IN THE HUMAN


GENOME
Gene Level
Sequence variation is the basis of functional variation
Affects directly the biochemical properties of proteins or,
indirectly, their regulation of expression

Genome Level
Phenotypes are the result of many genes acting in concert
Sexual recombination provides enormous potential of
diversity by combining a handful variants of genes

Population Level
Populations: are compartments of genotypic variation:
different populations have distinctive population genetic
profiles
Populations are natures testing ground for the fitness
of particular genotypic variants:
the environment of a population shapes its genetic profile

Monogenic disease

Complex disease

Mutation

Gene B

Gene

Genetic variations

Gene A

Gene C

Inheritance pattern
(dominan or recessive
Inheritance pattern
(complex)

Genetic risk in
population

Environmental/life-style

Genetic risk in population

Genetic risk in different families

Genetic risk in different families

GENOTYPE-PHENOTYPE
RELATIONSHIP
e
as
e
Dis

n
Cli

Mu

on
i
t
ta

Dy

c
n
u
sf

tion

Cell

Pathology
DNA

Molecular Genetics

lm
a
ic

t
s
e
f
i
an

Tissue

Biochemistry
Physiology

Family
Population

ns
o
i
at

Organ

Imaging

Individual

Clinical

Clinical
Epidemiology