GENE EXPRESSION:

•GENE =SEGMENT OF DNA = BLUEPRINT FOR A

SINGLE PROTEIN

•EACH CELL CONTAINS ALL GENES !!!

•WHEN A GENE IS EXPRESSED, THE PROTEIN IT

CODES FOR IS SYNTHESIZED (AT RIBOSOMES)

•NOT ALL GENES ARE EXPRESSED IN ALL CELLS (OF A

MULTICELLULAR ORGANISM) OR AT ALL TIMES

Gene expression

(EUCARYOTIC)(EUCARYOTIC)

DIFFERENTIATION

•DIFFERENT CELL TYPES SYNTHESIZE DIFFERENT PROTEINS•TYPICAL EUCARYOTIC CELL – ABOUT 10,000 PROTEINS•FOR THE 2000 MOST ABUNDANT (>50,000 COPIES/CELL)

FEW DIFFERENCES ARE FOUND (5 FOLD OR LESS)• A FEW % ARE CELL TYPE SPECIFIC

A PARTICULAR CELL’S EXPRESSED GENES VARY •WITH TIME•IN RESPONSE TO EXTERNAL SIGNALS•IN RESPONSE TO INTERNAL “CLOCKS”•DEPENDING ON STATE – NORMAL, STRESSED, ABNORMAL

HOW IS GENE EXPRESSION REGULATED??

TRANSCRIPTION

Transcription

M-RNA IS PROCESSED (SPLICED)

REGULATION AT DIFFERENT STEPS

KINDS OF REGULATION

M-RNA IS PROCESSED (SPLICED)

TRANSCRIPTIONAL CONTROL

RNA PROCESSING CONTROL

RNA TRANSPORT CONTROL

TRANSLATIONAL CONTROL

RNA DEGRADATION

PROTEIN ACTIVITY

PREVALENT (NO SYNTHESIS)

TRANSCRIPTION – CLOSER LOOK

RNA POLYMERASE ATTACHES TO DNA, MOVES ALONG IT, OPENS DOUBLE HELIX, SYNTHESIZES MRNA.

CONTROL OF EXPRESSION BY ASSISTING OR BLOCKINGATTACHMENT OF RNA POLYMERASE TO DNA.

RNA POLYMERASE BINDS AT A SPECIFIC REGION, THEPROMOTER, AT THE START OF THE GENE, IF AN ACTIVATOR IS ATTACHED AT AN ADJACENT SPECIFIC REGULATORY BINDING SEQUENCE (OPERATOR) AND AREPRESSOR IS NOT ATTACHED TO ITS OWN OPERATOR

TRANSCIPTION activator

TRANSCRIPTION START

6-2,6-4

TRANSCRIPTION – CLOSER LOOK

RNA POLYMERASE ATTACHES TO DNA, MOVES ALONG IT, OPENS DOUBLE HELIX, SYNTHESIZES MRNA.

CONTROL OF EXPRESSION BY ASSISTING OR BLOCKINGATTACHMENT OF RNA POLYMERASE TO DNA.

RNA POLYMERASE DOES NOT BIND AT THEPROMOTER, AT THE START OF THE GENE, IF AREPRESSOR IS ATTACHED TO ITS OWN OPERATOR

TRANSCIPTION repressor

TRANSCRIPTIONFACTORS

RECOGNITION OF BINDING MOTIFS IN DNA

RECOGNITION OF BINDING MOTIFS IN DNA 2

HOMEODOMAIN

TRYPTOPHAN REPRESSOR-SWITCH

OPERON

REGULATORY“NETWORK”

lac OPERON – E-COLI (PROCARYOTIC)

OPERON – SET OF GENES PLACED ONE AFTER THE OTHERON DNA, TAKING PART IN ONE PROCESS (BREAKDOWN OFlactose). E-COLI PREFERS glucose – WILL PROCESS lactose ONLY UNDER ( –glucose/+ lactose ) CONDITIONS. 4-SWITCH!

Cyclic AMP CONCENTRATION WHEN +glucose allolactose CONCENTRATION WHEN +lactose

ACTIVATOR: + (ACTIVE IF COMPLEX)

--glucose/+lactose

LAC OPERON KEEP!

REPRESSOR: + (ACTIVE IF FREE)

9 – 21 lac 4-way switch

EUCARYOTIC – MUCH MORE COMPLEX

• RNA POLYMERASE CANNOT INITIATE TRANSCRIPTION GENERAL TRANSCRIPTION FACTORS ASSEMBLE, FORM

COMPLEX ON OPERATOR NEAR PROMOTER - ABUNDANT

EUCARYOTIC 9 – 30,31

TBP – SUBUNIT OF TFIID

• SPECIFIC REGULATORS OF TRANSCRIPTION (ENHANCERS)

CAN ATTACH TO DNA MANY 1000 OF BP UPSTREAM,

CAN EVEN BE PLACED DOWNSTREAM FROM START SITE

VERY MINUTE AMOUNT PRESENT

• MAY NEED MORE THAN ONE TRANSCRIPTION FACTOR TO

ACTIVATE GENE

9-36,9-45

control of human beta-globin

proximity of GAL4enhances 1000 foldthe attachment of TFIIB to TFIID

REGULATORY NETWORKS

• TRANSCRIPTION FACTORS ARE PROTEINS THAT

ACTIVATE OR REPRESS GENES’ TRANSCRIPTION INTO

PROTEINS

• PROTEINS FORM COMPLEXES THAT INDUCE /TURN OFF

/REGULATE A GENE’S TRANSCRIPTIONAL CAPACITY

COMPLEX NETWORKS OF REGULATION OF GENE

EXPRESSION EMERGE

REGULATORY NETWORKS

HALLMARKS 1

CANCER IS CAUSED BY THE BREAKDOWN OF SEVERAL IMPORTANT NETWORKS,THAT GUARD AGAINST

UNCONTROLLED PROLIFERATION

NORMAL CELL STATES & CELL CYCLE

Check Points (Internal and External signals)

G1 –gap, decide whether to proliferate, wait

or cross to non-dividing stage G0

S -- DNA Synthesis

G2– gap, allow DNA repair

M – Mitosis, cell division

NORMAL ENTRY TO/EXIT FROM CELL CYCLE

Cell DivisionProliferation

Cell cycle arrest (G0) or

ProgrammedCell Death(Apoptosis)

Limited replication, senescence, crisis

Terminal differentiation

Check Point (Internal and External signals)

Growth Signals

Induced Apoptosis too many divisions

Anti Growth Signals

Cancer Cell-HALLMARKS

Cell DivisionProliferation

Cell cycle arrest or

ProgrammedCell Death(Apoptosis)

Limited replication, senescence, crisis

Terminal differentiation

1

3 3 4

2

Defective computation at check points, or failure to interpret signals or execute instructions:1 Proliferation becomes independent of growth factors.2 Loosing responses to cell cycle inhibitory signals.3 Failure to apoptose when necessary.4 Immortalization.

series of random genetic accidents

These are the main 4 HALLMARKS OF CANCER.

1. SELF SUFFICIENCY IN GROWTH SIGNALS

IN NORMAL CELLS, GROWTH FACTORS ARE RELEASED BY NEIGHBOR CELLS, BOUND BY GF RECEPTORS (USUALLYIN CELL MEMBRANE), WHICH GET MODIFIED AND INITIATE A CASCADE OF SIGNALING EVENTS .

1. Autonomous generation of growth factors2. Receptor overexpression or alteration3. Defective downstream processing

MUTANT Ras SEND DOWNSTREAM GROWTH SIGNALS WITHNO STIMULUS FROM UPSTREAM

What can go wrong?

2. IGNORING ANTI-GROWTH SIGNALS

IN NORMAL CELLS, MOST ANTIGROWTH SIGNALS DIRECTING THE CELL TO G0 ARE CHANNELED THROUGH THE Rb(RETINOBLASTOMA) PROTEIN.

TERMINAL DIFFERENTIATION IS INDUCED BY FORMATION OF Myc-Max COMPLEX.

3. EVADING APOPTOSIS

THE APOPTOTIC MACHINERY RELIES ON SENSORS (THAT DETECT INTERNAL AND EXTERNAL SIGNALS) AND EFFECTORS, THAT INDUCE AND CARRY OUT THE DEATHSENTENCE. p53 IS A CENTRAL PLAYER IN APOPTOSIS.

4. IMMORTALIZATION

A CELL CAN UNDERGO A LIMITED NUMBER OF DIVISIONS.

THE “COUNTING DEVICE” IS A STRING OF SEVERAL 1000

REPEATS OF A 6-BP SEQUENCE ELEMENT AT THE END OF

THE CHROMOSOMES (TELOMERS). IN EACH DIVISION 50 –

100 TELOMERIC BP ARE LOST. WHEN THEY RUN OUT, THE

CHROMOSOME ENDS ARE UNPROTECTED AND FUSE,

LEADING TO CRISIS AND DEATH OF THE CELL.

CANCER CELLS ACQUIRE TELOMERE MAINTENANCE

TWO MORE:

5. Formation of new blood vessels in the tumor (Sustained Angiogenesis) (NEEDED TO ACHIEVE LARGE SIZE)

6. Acquirement of metastatic behavior (cancer cells spread to vital organs) (CAUSE OF 90% OF CANCER RELATED DEATHS)