Gene Expression Regulation and Stem Cell Technology

Gene Expression Regulation

Nuclear Architecture and Gene Expression

The spatial organization of the nucleus, known as nuclear architecture, plays a crucial role in regulating gene expression. The arrangement of chromatin and the positioning of genes within the nucleus can influence whether genes are accessible for transcription.

• Nuclear architecture: The three-dimensional arrangement of chromatin and nuclear bodies within the nucleus.

• Gene expression: Genes located in tightly packed heterochromatin are generally less active, while those in loosely packed euchromatin are more likely to be transcribed.

• Example: Genes moved to the nuclear periphery may be silenced due to association with repressive nuclear lamina.

Alternative RNA Splicing

Alternative RNA splicing is a process by which different combinations of exons are joined together to produce multiple mRNA variants from a single gene, greatly increasing protein diversity.

• Key Point: Humans have about 20,000 genes but can produce over 100,000 proteins due to alternative splicing.

• Example: The troponin T gene can be spliced in different ways to produce muscle-specific proteins.

mRNA Degradation Signals

Nucleotide sequences that affect the degradation of mRNA are typically found in the untranslated regions (UTRs) of the mRNA, especially the 3' UTR.

• Key Point: These sequences determine the stability and lifespan of the mRNA, influencing how much protein is produced.

Blocking Translation Initiation

The initiation of translation can be blocked by regulatory proteins or microRNAs binding to the mRNA, preventing ribosome assembly and thus turning off gene expression.

• Key Point: This is a form of post-transcriptional regulation.

• Example: Iron regulatory proteins can block translation of ferritin mRNA in low iron conditions.

Ubiquitin and Proteasomes

Ubiquitin is a small protein that tags other proteins for degradation. Proteasomes are large protein complexes that recognize ubiquitin-tagged proteins and degrade them into peptides.

• Key Point: This system controls protein levels and removes damaged or misfolded proteins.

MicroRNAs (miRNAs) and Gene Silencing

MicroRNAs (miRNAs) are small non-coding RNAs that regulate gene expression post-transcriptionally.

• Formation: miRNAs are transcribed as primary miRNAs, processed into precursor miRNAs, and then into mature miRNAs by Dicer enzyme.

• Gene silencing mechanisms:

  1. miRNAs can bind to complementary sequences in mRNA, leading to mRNA degradation.

  2. miRNAs can inhibit translation of the target mRNA.

Determination and Differentiation

Determination is the process by which a cell becomes committed to a specific fate. Differentiation is the process by which a determined cell develops its specialized structure and function.

• 220 one-way streets: Refers to the approximately 220 different cell types in the human body, each following a unique developmental pathway.

Cytoplasmic Determinants and Induction

Gene expression in early development is influenced by cytoplasmic determinants (molecules in the egg cytoplasm) and induction (signals from neighboring cells).

• Cytoplasmic determinants: Unevenly distributed molecules that direct cell fate after fertilization.

• Induction: Cell-to-cell signaling that alters gene expression in target cells.

Microarray Analysis

Microarray analysis is a technique used to compare gene expression profiles between different cell types, such as normal and cancer cells.

• Steps:

  1. Isolate mRNA from both cell types.

  2. Convert mRNA to cDNA and label with fluorescent dyes (e.g., red for cancer, green for normal).

  3. Hybridize cDNA to a microarray chip containing thousands of gene probes.

  4. Scan the chip to detect fluorescence and analyze gene expression differences.

Microarray Color Interpretation

Color	Meaning
Red	Gene is expressed more in cancer cells
Green	Gene is expressed more in normal cells
Yellow	Gene is equally expressed in both cell types
Black	Gene is not expressed in either cell type

Stem Cells and Cloning

Stem Cells

Stem cells are undifferentiated cells capable of self-renewal and differentiation into specialized cell types.

• Embryonic stem cells: Pluripotent, can become any cell type in the body.

• Adult stem cells: Multipotent, limited to differentiating into a narrower range of cell types.

Somatic Cell Nuclear Transfer (Dolly the Sheep)

Somatic cell nuclear transfer (SCNT) is a cloning technique where the nucleus of a somatic cell is transferred into an enucleated egg cell.

• Steps:

  1. Remove the nucleus from an egg cell.

  2. Insert the nucleus from a donor somatic cell.

  3. Stimulate the egg to divide and develop into an embryo.

  4. Implant the embryo into a surrogate mother.

• Example: Dolly the sheep was the first mammal cloned using SCNT.

Induced Pluripotent Stem Cells (iPS Cells)

iPS cells are adult cells reprogrammed to a pluripotent state by introducing specific genes.

• Process:

  1. Introduce genes encoding transcription factors (e.g., Oct4, Sox2, Klf4, c-Myc) into adult cells.

  2. Cells revert to a pluripotent state, similar to embryonic stem cells.

• Application: iPS cells can be used for disease modeling, drug testing, and regenerative medicine.

Additional info: iPS cell technology avoids ethical issues associated with embryonic stem cells.