Molecular Biology of the Cell – Comprehensive Final Review

A broad set of question-and-answer flashcards covering key concepts, definitions, experimental methods, and examples from the Molecular Biology of the Cell study guide. Designed to aid comprehensive final exam preparation.

What is reverse genetics?

An approach that starts with an unknown DNA sequence and uses predictions and experiments to determine its function.

In gene mapping, what do the terms "upstream" and "downstream" correspond to?

They describe positions relative to transcription direction; upstream = 5' side, downstream = 3' side of the coding strand.

Which nucleotide is numbered +1 in a gene?

The first nucleotide transcribed into the primary RNA transcript.

Define an open reading frame (ORF).

A long stretch of codons without stop codons, predicting a protein-coding region.

What are introns?

Non-coding regions transcribed into pre-mRNA but removed by splicing before translation.

What are exons?

Sequences that remain in mature mRNA after splicing; include coding sequence and UTRs.

What is alternative splicing?

Production of multiple mRNA isoforms (and proteins) from one gene by varying exon inclusion.

Name two N-terminal targeting signals predictable from sequence similarity.

Signal peptide (ER/secretory pathway) and transit peptide (chloroplast import).

What is a cis-acting element?

A DNA sequence that regulates gene function on the same molecule (e.g., promoter, enhancer).

How do cis elements differ from trans-acting factors?

Trans-acting factors are typically proteins or RNAs that can diffuse and act on multiple DNA molecules.

Key difference between promoter and enhancer?

Promoter is adjacent to transcription start and recruits polymerase; enhancer can be distant and boosts promoter activity.

Monocistronic vs. polycistronic transcripts?

Monocistronic (eukaryotes) encode one ORF; polycistronic (prokaryotes) encode several ORFs in one mRNA.

What does sequence conservation imply?

High conservation suggests functional importance due to selective pressure against mutation.

Give two experimental ways to show that a gene is transcribed.

RNA blot (Northern) detection or isolation of a matching cDNA lacking introns.

How can promoter function be tested experimentally?

Fuse promoter to a reporter gene, reintroduce into cells, and measure reporter expression.

Purpose of a gene knock-out mutant?

To reveal the gene's role by observing phenotypic changes when the gene is inactivated.

What technique introduces specific nucleotide changes in a gene in vitro?

Site-directed mutagenesis.

PCR stands for?

Polymerase Chain Reaction.

What does a microarray (gene chip) measure?

Relative abundance of many RNA transcripts simultaneously via hybridization signals.

Name three classes of repetitive DNA elements.

SSRs (simple sequence repeats), SINEs, and LINEs.

What are pseudogenes?

Nonfunctional gene copies that accumulate mutations after gene duplication or retrotransposition.

Define nucleosome.

A structural unit of chromatin consisting of DNA wrapped around a histone octamer.

Euchromatin vs. heterochromatin?

Euchromatin is loosely packed and transcriptionally active; heterochromatin is tightly packed and generally silent.

Direction of DNA polymerase elongation?

5' → 3' using dNTP hydrolysis for energy.

Role of primase at the replication fork?

Synthesizes short RNA primers to provide 3'-OH ends for DNA polymerase.

What joins Okazaki fragments?

DNA ligase.

Function of telomerase?

Extends chromosome ends with repetitive sequences using its RNA template.

What damage does UV light cause in DNA?

Formation of thymine dimers.

Base-excision vs. nucleotide-excision repair?

Base excision removes one damaged base; nucleotide excision removes a short stretch of nucleotides including the lesion.

What is mismatch repair?

System that removes replication errors by distinguishing old and new DNA strands (often via methylation marks).

Translesion DNA synthesis is performed by?

Low-fidelity polymerases (e.g., DNA Pol V in E. coli) that bypass lesions like thymine dimers.

Homologous recombination occurs during which meiotic phase?

Prophase I (crossing-over).

Site-specific recombination in immune cells generates?

Diverse antibody genes (immunoglobulin gene rearrangement).

In prokaryotes, what sequence helps ribosomes find the start codon?

Shine-Dalgarno sequence.

Eukaryotic ribosome identifies the start codon by?

Scanning from the 5' cap to the first AUG in a Kozak consensus context.

First amino acid in prokaryotic translation?

Formyl-methionine (fMet).

Role of release factor in translation?

Recognizes stop codon and triggers polypeptide release and ribosome dissociation.

What is RNA interference (RNAi)?

Sequence-specific degradation or silencing of mRNAs by small RNAs (miRNA or siRNA) and RISC.

Chaperone vs. chaperonin?

Chaperones (e.g., HSP70) prevent premature folding; chaperonins (e.g., GroEL) provide isolation chambers for folding.

Disulfide bond forms between which amino acid residues?

Cysteines (oxidation of two –SH groups).

N-myristoylation adds what group and where?

A myristate fatty acid to the N-terminal glycine, replacing the start methionine.

Prenylation attaches isoprenoid groups to which residue?

Cysteine near the C-terminus (often in a CAAX motif).

O-linked glycosylation occurs where?

On serine or threonine side chains, mainly in the Golgi.

N-linked glycosylation occurs in which organelle?

Endoplasmic reticulum (ER) on asparagine residues.

What targets proteins for degradation by the proteasome?

Polyubiquitin tags.

Signal recognition particle (SRP) function?

Binds emerging signal peptide, pauses translation, and directs ribosome to ER membrane.

Stop-transfer sequence does what?

Triggers lateral release of a transmembrane segment, anchoring the protein in the membrane.

Import of nuclear proteins requires what signal?

Nuclear localization signal (NLS).

Role of Ran-GTP in nuclear transport?

Binds importin to release cargo in nucleus and drives directionality of import/export cycles.

Name the outer and inner membrane translocases of mitochondria.

TOM complex (outer) and TIM complex (inner).

Which complexes mediate chloroplast protein import?

TOC (outer) and TIC (inner) translocon complexes.

COPII-coated vesicles move in which direction?

Anterograde (ER → Golgi).

COPI-coated vesicles are generally involved in?

Retrograde transport (Golgi → ER or within Golgi).

Clathrin primarily coats vesicles budding from which region?

Trans-Golgi network and plasma membrane.

ARF1-GTP initiates what process?

Recruitment of adaptor proteins and coat assembly for vesicle formation.

SNARE proteins function in vesicle trafficking how?

v-SNAREs on vesicles and t-SNAREs on targets pair to drive membrane fusion.

Rab GTPases provide what during vesicle docking?

Molecular identity and regulation, recruiting tethering factors to target membranes.

Define anterograde traffic.

Forward movement of vesicles from ER through Golgi toward plasma membrane or lysosome.

Define retrograde traffic.

Backward vesicle movement returning escaped or resident proteins to earlier compartments.

What is the ERGIC?

ER-Golgi intermediate compartment formed by fusion of COPII vesicles before Golgi entry.

Function of KDEL sequence.

C-terminal retrieval signal that returns ER-resident soluble proteins from Golgi via COPI vesicles.

Microarray spot color indicates what?

Relative hybridization of labeled sample RNAs: e.g., red vs. green intensities show expression differences.

SSR stands for?

Simple Sequence Repeat (tandem short repeat DNA).

Difference between LINEs and SINEs?

Both are retrotransposons; LINEs are long and often encode reverse transcriptase, SINEs are short and non-coding.

What is a processed pseudogene?

A gene copy derived from reverse-transcribed mRNA inserted back into the genome, lacking introns and promoter.

Telomere repeats are added using what template?

An RNA sequence carried within telomerase.

Photoreactivation repairs what lesion?

UV-induced thymine dimers via photolyase and visible light.

What triggers transcription termination in prokaryotes?

Hairpin stem-loop followed by poly-U tract in the RNA or Rho-dependent mechanisms.

Role of sigma factor in bacteria?

Confers promoter specificity to RNA polymerase during initiation.

Mediator complex assists transcription by?

Bridging enhancer-bound activators with the promoter-bound pre-initiation complex.

Negative elongation factors cause?

Pause of RNA Pol II ~50 nt downstream of initiation until released by signals.

Histone H3 modifications influence what?

Chromatin packing and thus accessibility of DNA to transcription machinery.

5' cap structure?

7-methylguanosine linked via an unusual 5'-5' triphosphate bond.

Purpose of poly(A) tail.

Enhances mRNA stability, nuclear export, and translation initiation.

Spliceosome recognizes intron boundaries using what RNA component?

U1 snRNA (binds 5' splice site) and others (U2, U4/U6, U5).

Exon Junction Complex (EJC) marks what?

Sites of splicing; assists in nonsense-mediated decay if not removed during translation.

Shine-Dalgarno sequence pairs with which rRNA?

16S rRNA of the prokaryotic small ribosomal subunit.

Kozak consensus surrounds which codon?

The start codon (AUG) in eukaryotic mRNAs, enhancing initiation efficiency.

During elongation, peptide bond formation transfers the polypeptide from which site to which?

From the P-site tRNA to the A-site tRNA.

Polysome definition?

A single mRNA simultaneously being translated by multiple ribosomes.

Heat-shock proteins (HSP70) role during import?

Keep polypeptides unfolded for translocation into organelles such as mitochondria or chloroplasts.

Function of glycosyl-phosphatidylinositol (GPI) anchor.

Attaches proteins to the outer leaflet of the plasma membrane after ER/Golgi processing.

Signal peptide is typically removed by what enzyme?

Signal peptidase in the ER lumen.

CAH1-GFP fusion helped demonstrate what about CAH1?

That CAH1 travels through the secretory pathway and localizes to chloroplasts.

Brefeldin A (BFA) blocks what step in secretion?

Vesicle transport from ER to Golgi, causing cargo accumulation in ER/Golgi and loss from final destination.

Cycloheximide effect used in Villarejo et al.?

Inhibits new protein synthesis to track relocation of existing CAH1-GFP after BFA washout.

Why does Villarejo lane 4 (CAH without SP + microsomes) show no glycosylation?

Signal peptide is required for ER import; without it the protein stays cytosolic and bypasses ER-based glycosylation.

Central dogma stages affected by gene features?

DNA → RNA (transcription), RNA processing, RNA → protein (translation), and post-translational events.