Pro and Eukaryotic 13.1-13.3

Prokaryotic Regulation

Bacteria do not require the same enzymes all the time.

Enzymes are produced as needed.

François Jacob and Jacques Monod (1961) proposed the operon model to explain regulation of gene expression in prokaryotes.

•An operon is a group of structural and regulatory genes that function as a single unit.

Operon Components

A regulatory gene  that codes for a repressor protein controls the operon.

•Regulatory gene is normally located outside the operon.

•Repressor protein controls whether the operon is active or not.

Promoter: DNA sequence where RNA polymerase first attaches. Short segment of DNA.

Operator: DNA sequence where active repressor binds. Short segment of DNA.

Structural Genes: One to several genes coding for enzymes of a metabolic pathway. Transcribed simultaneously as a block. Long segment of DNA.

The trp Operon

The regulator codes for a repressor.

If tryptophan (an amino acid) is absent:

•The repressor is unable to attach to the operator (expression is normally “on”).

•RNA polymerase binds to the promoter.

•Enzymes for synthesis of tryptophan are produced.

If tryptophan is present: It combines with the repressor protein as its corepressor. Repressor becomes functional when bound to tryptophan. Repressor blocks synthesis of enzymes in the pathway for tryptophan synthesis.

The lac Operon 1

The regulator codes for a repressor.

If lactose (a sugar that can be used for food) is absent: The repressor attaches to the operator. The expression is normally “off.”

If lactose is present: It combines with the repressor and renders it unable to bind to the operator. RNA polymerase binds to the promoter. The three enzymes necessary for lactose catabolism are produced.

Further Control of the lac Operon 1

E. coli preferentially breaks down glucose. The lac operon is maximally activated only in the absence of glucose.

When glucose is absent:

•Cyclic AMP (cAMP) accumulates.

•cAMP binds to catabolite activator protein (CAP).

•CAP, when bound to cAMP, binds to a site near the lac promoter.

•When CAP is bound, RNA polymerase binds better to the promoter.

•The structural genes of the lac operon are expressed more efficiently.

Further Control of the lac Operon 2

When glucose is present:

•There is little cAMP in the cell.

•CAP is inactive.

The lac operon is not expressed maximally.

Eukaryotic Regulation

A variety of mechanisms. Five primary levels of control:

Nuclear levels:

•Chromatin structure.

•Transcriptional control.

•Posttranscriptional control.

Cytoplasmic levels:

•Translational control.

•Posttranslational control.

Chromatin Structure 1

Eukaryotic DNA is associated with histone proteins.

•Together they make up chromatin.

Nucleosomes.

•DNA is wound around groups of eight molecules of histone proteins.

•It looks like beads on a string.

•Each bead is called a nucleosome.

The levels of chromatin packing are determined by the degree of nucleosome coiling.

Chromatin Structure 2

Euchromatin

•Loosely coiled DNA

•Transcriptionally active

Heterochromatin

•Tightly packed DNA

•Transcriptionally inactive

Barr Body

Females have two X chromosomes, but only one is active. The other X chromosome is tightly packed along its entire length and is inactive. The inactive X chromosome is called a Barr body.

•An example is a female calico cat, which has patches of orange and black in its coat, depending on which X chromosome is in the Barr bodies of cells in patches.

Epigenetic Inheritance

Histone modifications may be linked to epigenetic inheritance, in which variations in the pattern of inheritance are not due to changes in the DNA nucleotide sequence.

Epigenetic inheritance refers to inheritance patterns not dependent on genes themselves.

Explains unusual inheritance patterns It may also play a role in growth, aging, and cancer.

Transcriptional Control

Transcription is controlled by proteins called transcription factors.

Transcription factors are proteins that help regulate transcription by assisting RNA polymerase in binding to the promoter.

A transcriptional activator is a DNA-binding protein that:

•Binds to enhancer DNA.

•Regions of DNA where factors that regulate transcription can also bind.

•Transcription factors are always present in the cell, but most likely have to be activated before they will bind to DNA.

Posttranscriptional Control

Posttranscriptional control operates on the primary mRNA transcript.

Given a specific primary transcript, intron excision can vary. Splicing of exons can vary. It determines the type of mature transcript that leaves the nucleus.

May also control the speed of mRNA transport from the nucleus
to the cytoplasm.

•It will affect the number of transcripts arriving at the rough ER and, therefore, the amount of gene product realized per unit time.

Ex: The hypothalamus and thyroid gland produce a protein hormone called calcitonin, but the mRNA that leaves the nucleus is not the same in both types of cells.

•The thyroid and hypothalamus, therefore, release different versions of the hormone.

Small RNA (sRNA) Molecules Regulate Gene Expression

The noncoding transcribed DNA is used to form small RNA (sRNA) molecules.

•These are involved in gene regulation and function at multiple levels of gene expression.

•Some of the sRNA molecules regulate transcription and others regulate translation.

•sRNA molecules are the source of microRNAs (miRNAs) regulating translation by causing the destruction of mRNAs before they can be translated.

•They also are the source of small-interfering RNAs (siRNAs) that form a silencing complex that targets specific mRNAs for breakdown, preventing their expression.

This is referred to as RNA interference

Translational Control

Translational control determines the degree to which mRNA is translated into a protein product. Features of the mRNA affect whether translation occurs and how long the mRNA remains active.

•Presence of 5′ cap.

Length of poly-A tail on 3′ end.

Posttranslational Control

Posttranslational control affects the activity of a protein product.

Posttranslational control is accomplished by regulating.

•Activation

•Degradation rate

Proteases

These enzymes break down proteins, thereby helping regulate gene expression.

•They regulate how long a protein remains active in the cell

•They are confined to proteasomes or lysosomes to protect the cell

Gene Mutations

A gene mutation is a permanent change in the sequence of bases in DNA. The effects of a gene mutation can range from

•No effect on protein activity.

•Complete inactivation of the protein.

Germ-line mutations occur in sex cells. Somatic mutations occur in body cells.

Causes of Mutations

Spontaneous mutations

Chemical changes in DNA that lead to mispairing during replication. Movement of transposons from one chromosomal location to another.

Replication Errors.

•DNA polymerase.

•Proofreads new strands

•Generally corrects errors

•Overall mutation rate is 1 in 1,000,000,000 nucleotide pairs replicated.

Induced mutations

Caused by mutagens such as radiation and organic chemicals. Many mutagens are also carcinogens (cancer-causing).

Environmental Mutagens.

•Food.

Tobacco smoke.

Effect of Mutations on Protein Activity

Point mutations: One type of point mutation is a base substitution. Involves a change in a single DNA nucleotide. Change one codon to a different codon.

Effects on the protein vary:

•Nonfunctional

•Reduced functionality

•Unaffected

Frameshift mutations: One or two nucleotides are either inserted or deleted from DNA. The protein is always rendered nonfunctional.

•Normal: THE CAT ATE THE RAT

•After deletion: THE ATA TET HER AT

After insertion:THE CCA TAT ETH ERA T

Nonfunctional Proteins

If a faulty enzyme is inserted into a metabolic pathway, a person may be unable to convert one molecule to another with serious consequences.

In the disorder phenylketonuria (PKU), phenylalanine builds up in the system and the excess causes an intellectual disability.

In androgen insensitivity, cells are unable to respond to testosterone because of a defective androgen receptor.

•Female instead of male genital form, and female instead of male secondary sex characteristics occur at puberty.

Mutations Can Cause Cancer

The development of cancer involves a series of accumulating mutations. Proto-oncogenes – Stimulate cell division

•Mutated proto-oncogenes become oncogenes that are always active.

Tumor suppressor genes – inhibit cell division.

Mutations in oncogene and tumor suppressor genes:

•Stimulate the cell cycle uncontrollably.

This leads to tumor formation.