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RPI / Biology / BIOL 1010 / What is a model organism to understand prokaryotic gene expression?

What is a model organism to understand prokaryotic gene expression?

What is a model organism to understand prokaryotic gene expression?

Description

School: Rensselaer Polytechnic Institute
Department: Biology
Course: Introduction to Biology
Term: Spring 2016
Tags: Biology
Cost: 25
Name: Week 9 Notes- Bio
Description: Molecular Biology 3 Emerging Diseasees
Uploaded: 04/03/2016
53 Pages 77 Views 1 Unlocks
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Control of Gene Expression


What is a model organism to understand prokaryotic gene expression?



Eukaryotic cell

Animation: Chime YouTube Version Windows media (get plugin)


Why lactose is not always present in the diet?



Gene Expression: The process by which information flows  from DNA to RNA to protein or genotype to phenotype.

E. coli as model organism to understand prokaryotic gene  expression.

Features of prokaryotic  

transcriptional regulation

•Ability to change metabolic activities

in response to the environment.

•Capability to induce/repress entire  


Where does insulin synthesized?



enzyme pathways- genome  If you want to learn more check out What are two species of elephants?
Don't forget about the age old question of They are the building blocks of proteins. what are they?

organization.

•Examples of regulated gene  

networks: lac operon, trp operon

Lac operon: Proteins interacting with DNA turn prokaryotic  genes on & off. First regulation discovered by François  Jacob and Jacques Monod in 1961 We also discuss several other topics like Acidophiles grow best below at what ph?

• Lactose is a disaccharide that can be  

broken down into two monosaccharides  

used in glycolytic pathway

•Lactose is not always present in the diet.

•If present, three enzymes are needed to  

convert it to a useful metabolite

•The genes for the enzymes are physically  

linked on the genome

•Regulate transcription based upon  

presence or absence of lactose.

•Microbes are efficient!!

Lactose operon is not transcribed in the absence of  lactose.

3 enzymes located together on the chromosome and regulated  as single unit.  

β-galatosidase-lacZ

galactose permease-lacY

thiogalatoside transacetylase-LacA

Regulatory regions precede the genes encoding the enzymesWe also discuss several other topics like What is baroque?

Lac operon: alternative states depending upon presence  or absence of lactose. Model of induction.

Don't forget about the age old question of Why does the battle of kursk july-august 1943 matter?

•Lactose present– operon is active and available for transcription  because lactose binds to the repressor protein inducing a  conformational change We also discuss several other topics like What is the second step inflow of genetic information?

•Repressor•Lactose complex cannot bind to operator and RNA  polymerase binds & initiates transcription.

Attendance Exercise

There is a mutation such that the nucleotide  sequence of the operator is changed. The lac  repressor protein will not bind.

a. Explain what happens to transcription of the  lac operon when lactose is present? b. Explain what happens to transcription of the  lac operon when lactose is absent?

Unifying theme: Transcription of many prokaryotic genes  is up regulated as a group or turned off as a group. However,  there are alternatives to the lac example of induction.

•trp operon repressor requires binding of tryptophan to the  repressor protein for repression of transcription. •There are also operons controlled by activators which assist  RNA polymerase binding.

Eukaryotic mechanisms  

of gene regulation

1.All cells contain the same genotype. Differential gene expression leads to  different cell types.

2.Multiple mechanisms to regulate gene expression.

•DNA packing. Prevents access of the DNA for transcription. •Eukaryotic transcription has multiple enhancers, silencers, and variety  of proteins that are assembled into a complex.

•Alternative RNA splicing leads to multiple mRNAs for translation. •microRNAs and RNA interference.

•Translation and later events also regulated.  

Differentiation in cell types in multicellular  

organisms is due to differential gene expression

DNA packing prevents access of the DNA for transcription.

X-chromosome inactivation: In female mammals, one  X chromosome is inactivated in all cells except  gametes.

•Occurs early in embryonic development.

•Random which X is inactivated.

•Inactive X referred to as Barr body.

Gene expression during  embryogenesis

Video: NOVA  

Life’s Greatest Miracle  

chapter 3

(on the bottom part of the video pop

up, mouse over the slider and select  

episode 3)

“The embryo Takes 

Shape”

youtube copy backup link

Eukaryotic transcription complexes have multiple  proteins and regulatory factors .

Regulation of gene expression for development.

•Important first studies from Drosophila melanogaster.

•Identify mutants that developed abnormally. •Normal head with eyes and antenna shown on the left. •Abnormal development on the right with legs in place of  antenna

Early development of head-tail polarity. protein gradients

Post transcriptional regulation: Alternative RNA  splicing results in multiple transcripts from the same  gene.

Standard splicing to remove introns

Example of alternative splicing

Role of microRNAs (miRNA) in gene  expression.

•Discovered 1993

•Small single-stranded RNA  

(20-26 nucleotides long)

•Associate with protein  

complex and complementary  

mRNA sequence

•Imperfect base pairing is  

allowed

•Estimated that expression of  

1/3 of human genes are

regulated by ~1000 miRNAs

Gene expression regulated at later stages.

•mRNA degradation: t1/2 of transcripts variable. prokaryotic: very short, minutes, because mRNA  lasting for too long could have negative impact on  environmental change.

eukaryotic: variable, hours to months and can be  regulated particularly by hormones

•translation highly regulated

inhibitory protein blocks transcription of  hemoglobin mRNA if heme supply is low (iron  containing compund that binds oxygen)

Post-translational Regulation  Protein Activation

example Insulin activation

•Insulin synthesized in pancreas. •Proinsulin is inactive.

Multiple levels of regulation  in Eukaryotic cells

•In the nucleus & in cytoplasm. •Before & after translation. •Novel regulatory mechanisms are still being discovered.

DNA Microarrays

picomoles DNA

Emerging Infectious Diseases (EID). Where do they  originate? What do we do? Why do we care?

Global richness map of the geographic origins of EID events from 1940 – 2004. http://www.ucsd.tv/search-details.aspx?showID=23137

The Realities in Global Trends

•EIDs are a significant burden on global economies &  public health.

•EID emergence thought to be driven by socio-economic environmental & ecological factors.

http://www.prweb.com/releases/2005/03/prweb220610.htm

Definition of a Pandemic: Tracking  Swine Flu using Google Maps

http://www.gadgetlite.com/2009/04/27/tracking-swine-flu-pandemic-google/

pandemic – occurring over wide geographical  area and affecting an exceptionally high  proportion of the population.

zoonotic diseases or zoonosis: infectious  diseases that can be transmitted from other vertebrate  animals to humans. May be direct or via a vector.

http://www.avma.org/onehealth/appendix_c.asp

Zoonotic Diseases: Five Stages through which pathogens  of animals evolve to cause disease in humans.

Critical transitions:

•Stage 1 to Stage 2:  from animal to human

•Stages 3 & 4:  

pathogen’s ability to  sustain cycles of human  to human transmissions Ex. modern Ebola  outbreaks are stage 3.

2007 Nature 447, 279-283.

Global Trends Results

•EIDs events have risen significantly over time.

•Peak 1980s attributed to HIV.

•EIDs dominated by zoonoses (60.3%).

•71.8% of zoonoses originate in  

wildlife (SARs, Ebola) and this  

trend is increasing.

•54.3% caused by bacteria or  

rickettsia.

•25.4% by viruses or prions.

•Drug-resistant microbes are

Increasing.

•Data reveal substantial risk

of zoonotic & vector-borne EIDs  

at lower altitudes (below equator)

where reporting effort low.

2008 Nature 451, 990-994

List of NIAID Emerging and Re-emerging Diseases

Group I—Pathogens Newly Recognized in the Past Two  

Decades

Acanthamebiasis

Australian bat lyssavirus

Babesia, atypical

Bartonella henselae

Ehrlichiosis

Encephalitozoon cuniculi Encephalitozoon hellem Enterocytozoon bieneusi Helicobacter pylori

Hendra or equine morbilli virus Hepatitis C

Hepatitis E

Human herpesvirus 8

Human herpesvirus 6

Lyme borreliosis

Parvovirus B19

Group II—Re-emerging  Pathogens

Enterovirus 71

Clostridium difficile

Mumps virus

Streptococcus, Group A Staphylococcus aureus

Why are many EIDs linked to Viruses?

Viruses have

•High rate of mutation, contact between species, spread from  isolated populations.

•RNA viruses – unusually high rates of mutation

lack the proofreading mechanisms seen in DNA replication

•Examples of RNA virus EIDs: common cold, measles, mumps, polio HIV or AIDs  

Ebola – hemorrhagic fever

West Nile & other emerging viruses causing encephalitis SARS – severe acute respiratory syndrome (2002-2003)

•Examples of DNA virus EIDs:  

hepatitis, chicken pox, herpes infections

Ebola virus

Infectious Disease Cycle  & Persistence

Outbreak

Spread

Resistance

Dependent on duration of  infectivity in host.

•Rate of infection of new hosts. •Rate of development of host  protective immunity

•Population density, size, and  structure: may eradicate  locally but persists regionally.

How does an Influenza Pandemic  Occur?

Video youtube copy

Experts predict next epidemic will start in animals. USA Today. October 22, 2008.

First known case Swine Flu (H1N1): March 28, 2009.

First known case US: April 17, 2009

April 26th: New Zealand, France, Israel, Brazil, Spain report cases. 86 deaths in Mexico attributed to swine flu.

June 11th: Pandemic declared by WHO. The first influenza pandemic in the  last 40 years.

July 14th: WHO authorizes pharmacy companies to manufacture vaccines. August 27th: US colleges see spike in number of cases.

September 4th: WHO announced 625 deaths in the last week. October 5th: vaccine ready.

October 24: President declares a national emergency.

Attendance exercise:

Why did the President declare a national  emergency concerning swine flu?

What is the greatest fear of scientists and policy  

makers?

Swine Flu or H1N1 is an influenza A virus containing 11 viral genes.

Genome consists of

8 segments of single  

stranded RNA.

Genes encoded include:  

polymerase PB2

polymerase PB1& PB1-F2

polymerase PA

hemagglutinin HA

nuclear protein NP

neuraminidase NA

matrix proteins M1 & M2

Neumann et al. 2009. Nature 459, 931-939.

nonstructural proteins NS1 & 2

Schematic diagram of the influenza  

viral life cycle.

Animation: Influenza 

Virus Replication

Figure 2: G Neumann et al.

Nature 459, 931-939 (2009)  

Where did most recent H1N1 come from?

Original reservoir was birds

1918: human influenza A  

(H1N1) pandemic – transfer  

from chickens  

1918: Cedar Rapids Swine  

Show influenza A transmitted to  

pigs from humans.

1931: documented infectious  

transmission

1933: used ferret model to  

document transmissibility for  

human & swine viruses.

1918-present: evolution of  

virus in humans.

1957: H1N1 abruptly  

disappeared from humans.  

Morens et al. 2009. NEJM 361. 225-229.

Genesis of 2009 Swine-origin H1N1 Influenza Viruses.

Neumann et al. 2009. Nature 459, 931-939.

Monitoring for Influenza viruses MSNBC Video: H1N1 Virus We can watch virus evolution.

This requires large-scale sequencing of viruses isolated from  patients.

After this, bioinformatic analysis shows which ones are evolving  most rapidly. These are, on evolutionary principles, the most  likely to be problematic in the future.

video backup link 

US Vaccine Evaluation Centers

Prevention and Control

Possibility #1

Prevention and Control Possibility #2

Antiviral drugs interfere with viral  specific proteins:

adamantanes: block ion channel  formed by M proteins (M proteins  used for escape from  

endosome).

Oseltamivir (Tamiflu) and  aznamivir: neuraminidase  inhibitors block release of new  virus particles.

A

B

M protein with 2  adamantanes bound

Attendance exercise:

Why did the President declare a national  emergency concerning swine flu?

What is the greatest fear of scientists and policy  

makers?

Dengue Fever on the watch list. Why?

Video from ABC News 

(copy of the Video link) 

•Caused by one of 4 different, but related viruses. •Carried by mosquito, Aedes aegypti

•4-6 days for symptoms to appear.

• Often referred to as break-back fever

+

Why?

Local copy of the video

Antibiotic resistance: MRSA Infections

•Methicillin-resistant Staphylococcus aureus.

•Resistant to broad range of antibiotics.

•Harmless unless enters body through cut or wound. •Deadly to those with weakened immune systems. •Hospitals, assisted living facilities.

•Teams, locker rooms.

•2M people/year infected in hospitals. ~100,000 deaths.

Kellen Winslow, Oct 21, 2008

MRSA Infections – RPI News: August 16, 2010

•Develop antimicrobial coatings for hospital surfaces •Carbon nanotube-enzyme conjugates made with Lysostaphin, a cell wall degrading enzyme

•Lysostaphin from non-pathogenic strains of Staph bacteria had >99% MRSA killed within 2 hours.

R Pangule et al. JS Dordick

2010 ACS Nano 4, 3993-4000.

Antibiotics: An Uphill Battle

www.CEN-ONLINE.org April 14, 2008

•Short period of effectiveness.  

•Emerging drug-resistant pathogens.

•Cost of R&D without promise of profit margin. •Mother nature more clever than humans!

•Focus on natural products- novel compounds

The Need for New Ideas

Focus on Under Developed Nations

•Focus on Tuberculosis- second most lethal infectious disease •Bill and Melinda Gates Foundation – October 21, 2008

•X PRIZE to fight tuberculosis worldwide-effective diagnosis

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