Chapter 15 Micro

Origin Recognition Complex

• marks eukaryotic replication origins

CDC6/CDT1

• loading factors that help recruit MCM helicase during origin licensing

Mini-Chromosome Maintenance Complex (MCM)

• core unwinding motor in eukaryotic replication

CMG Complex

• active eukaryotic helicase complex composed of CDC45, MCM, and GINS

Replication protein a (RPA)

• eukaryotic single-stranded DNA-binding protein

Proliferating cell nuclear antigen (PCNA)

• the eukaryotic sliding clamp that increases polymerase processivity

Replication factor c (RFC)

• clamp loader for PCNA

Flap endonuclease 1( FEN1)

RTEL1

• helicase associated with telomere maintenance and replication termination functions

TATA-binding proteins (TBP)

• transcription factor that helped position RNA polymerase at promoters

TFIID

• general transcription factor complex that contains TBP

Pol I

• rRNA

Pol II

• mRNA

Pol III

• tRNA plus 5s rRNA

Pre-mRNA

• primary transcript containing both exons and introns before processing

5’ cap

• 7-methylguanosine added to the 5’ end of eukaryotic mRNA

Poly-A tail

• adenine rich 3’ tail that protects mRNA and supports export and translation

Spliceosome

• ribonucleoprotein complex that removes introns from pre-mRNA

Poly A binding protein (PABP)

• binds the 3’ tail and helps bridge mRNA during eukaryotic initiation

Shine-Dalgarno sequence

• bacterial ribosome binding site that aligns the start codon with 16s rRNA

Leaderless transcript

• mRNA lacking a long 5’ leader

• common in some archeal transcripts

Sec system

• conserved protein translocation system found across the 3 domains

Tat system

• twin-arginine translocation pathway used for certain protein targeting, especially in organelles

Heat shock proteins (HSP)

• chaperone involved in protein folding or refolding

Bacterial Genome structure

• one circular chromosome

Bacterial and Histones

• contain no true histones

• contains nucleoid associated proteins

Bacterial replication origins

• one origin

Bacterial replicative polymerase

• DNA polymerase III is the main replicative enzyme

Bacterial RNA polymerase

• single RNA polymerase with sigma factor

Bacterial mRNA organization

• often polycistronic

Bacteria and Introns

• rare

Bacterial transition initiation

• uses shine-dalgarno

• fMet initiator tRNA

Bacterial protein localization

• Sec plus vesicular trafficking

• direct organelle translocation

Eukaryotic genome structure

• multiple linear chromosomes

Eukaryotes and Histones

• DNA wrapped around histones in nucelosomes

Eukaryotic replication origins

• multiple origins per chromosomes

• chromosome are composed of many replicons

Eukaryotic replicative polymerases

• Pol α-primase initiates

• Pol ε leads

• Pol δ lags

Eukaryotic RNA polymerase

• Pol I, Pol II, Pol III

Eukaryotic mRNA organization

• usually monocistronic

Eukaryotic introns

• common in many genes

• removed by splicing

Eukaryotic translation initiation

• uses 5’ cap recognition

• scanning

• Met initiator tRNA

Eukaryotic protein localization

• Sec plus vesicular trafficking and direct organelle translocation

Why is compartmentation especially important in eukaryotic cells?

• eukaryotic cells are larger and more internally complex

• they need compartmentation to segregate reactions, concentrate molecules, and coordinate genome replication and expression efficiently.

What is the difference between a membrane-bound organelle and a biomolecular condensate?

• Membrane-bound organelles separate reactions with lipid membranes,

• biomolecular condensates separate and concentrate reactions through protein–RNA or protein–protein assemblies without a membrane.

Why do eukaryotic chromosomes require multiple origins of replication?

• eukaryotic chromosomes are very large and linear

• using many origins allows replication to proceed simultaneously at multiple sites and finish within a practical cell-cycle time frame.

Why is PCNA so important during eukaryotic DNA replication?

• PCNA is the eukaryotic sliding clamp

• It tethers polymerase to DNA, increases processivity

• especially important for efficient lagging-strand synthesis by Pol δ.

What is meant by origin licensing in eukaryotic DNA replication?

• Origin licensing is the loading of pre-replication machinery (especially the inactive MCM helicase) onto origins during late M or early G1 so those origins are permitted to fire in S phase.

Why must MCM helicase remain inactive when first loaded onto DNA?

• if MCM were active immediately, DNA strands could unwind too early, exposing them to damage or unscheduled replication

• Keeping it inactive until S phase preserves timing and genome integrity.

What roles do RPA and topoisomerases play during initiation?

• RPA coats single-stranded DNA to stop reannealing and secondary structure formation

• Topoisomerases relieve positive supercoils that build ahead of the advancing helicase.

Why does the lagging strand require repeated priming?

• DNA polymerases synthesize only 5′ to 3′

• the lagging-strand template must be copied discontinuously as a series of short fragments, each of which needs its own RNA primer.

What is the distinct role of Pol α compared with Pol δ and Pol ε?

• Pol α has low processivity and mainly initiates synthesis by extending the RNA primer

• Pol δ and Pol ε are high-processivity polymerases that perform most strand elongation.

How are RNA primers removed from Okazaki fragments in eukaryotes?

• Pol δ displaces primer RNA to create a short 5′ flap

• FEN1 cleaves the flap

• Repeated rounds of this process remove the primer so DNA ligase can seal the nick.

What is the end-replication problem?

• the final RNA primer at the 5′ end of the lagging strand is removed

• there is no upstream 3′-OH for polymerase to fill the gap

• the daughter chromosome becomes shorter after each round of replication.

How does telomerase solve this problem?

• Telomerase carries an internal RNA template and functions as a reverse transcriptase to extend the telomeric G-tail

• This creates enough extra sequence for conventional primase and polymerase to finish lagging-strand synthesis.

Why can inappropriate telomerase activation be dangerous?

• If telomerase is activated in cells that should normally senesce, those cells may keep dividing beyond normal limits

• can support malignant transformation.

Why are archaea often described as having bacterial-like genomes but eukaryotic-like information machinery?

• Many archaea have relatively small circular chromosomes like bacteria

• Key proteins involved in DNA replication and transcription are more closely related to eukaryotic systems.

What is recombination-driven DNA replication in archaea?

• a mechanism in which homologous recombination intermediates help trigger replication initiation

• provide an alternative route beyond standard origin firing.

Why does eukaryotic transcription require more processing than bacterial transcription?

• Eukaryotic transcripts are produced in the nucleus as pre-mRNAs that contain introns and therefore must be capped, spliced, and polyadenylated before export and translation

• Bacterial mRNAs are generally used more directly.

What is the functional significance of the 5′ cap and 3′ poly-A tail?

• 5′ cap and 3′ poly-A tail protect mRNA from degradation, mark it for export, and help it engage the translation machinery

• the 5’ cap also helps ribosomes recognize the transcript in the cytoplasm.

How are archaeal transcription systems both bacterial-like and eukaryotic-like?

• Archaeal transcription occurs in a non-nuclear, bacterial-like setting and often produces polycistronic mRNA

• the RNA polymerase and promoter recognition factors resemble eukaryotic Pol II systems.

What is the major difference between bacterial and eukaryotic translation initiation?

• Bacteria position the ribosome by Shine–Dalgarno pairing

• Eukaryotes recruit the small subunit to the 5′ cap and scan along the mRNA until the start codon is found.

Why is initiator tRNA chemistry a useful domain comparison?

• Bacteria begin translation with N-formylmethionine-tRNA

• Eukaryotes and Archaea use methionine initiator tRNA without formylation, helping distinguish their initiation strategies.

What is the purpose of the cap–tail bridge in eukaryotic translation?

• The bridge links the 5′ cap and 3′ poly-A tail through associated proteins

• helps to activate the mRNA for efficient ribosome recruitment and scanning.

Why do eukaryotes rely more heavily on protein-targeting systems than bacteria?

• Eukaryotic cells contain many membrane-bound compartments

• newly made proteins must often be sorted and transported to the ER, mitochondria, chloroplasts, Golgi, lysosomes, or plasma membrane.

What is the general role of a signal peptide?

• A signal peptide is an amino-terminal targeting sequence that directs a newly synthesized protein to a translocation pathway such as the Sec system.

How do vesicular transport and direct translocation differ?

• Vesicular transport moves proteins inside membrane-bound carriers between compartments

• Direct translocation threads a protein through a translocon across a membrane.

Why is chromatin remodeling essential for eukaryotic gene expression?

• DNA is wrapped around nucleosomes

• Transcription factors and polymerases cannot efficiently reach promoter sequences unless chromatin structure is repositioned or opened.