MHAM

COLLEGE OF MEDICINECLASS 2024

Genome
Jan Michael Lao, RMT, M.D. | August 13, 2021
GENERAL PATHOLOGY

Outline a. Micro-RNAs
b. Long non-coding RNAs
I. The Human Genome 3. Epigenetics
A. Noncoding DNA → heritable changes in gene expression that
are not caused by alterations in DNA
B. Noncoding RNA
sequence
C. Epigenetics → histone modification can be done through
II. Gene Editing acetylation, methylation, ubiquitination,
citrullination and phosphorylation of
I. THE HUMAN GENOME specific amino acids within the histone
A. Noncoding DNA protein
→ Chromosomes
▪ where DNAs are contained • Non-coding DNA
▪ composed of chromatids → Genetic polymorphisms
→ Heterochromatin a. Single nucleotide polymorphisms (SNPs)
▪ densely packed, inactive ▪ variants at a single nucleotide position
▪ unwound form of chromatids ▪ occur across the genome within exons,
→ Euchromatin introns, and intergenic regions
▪ disperse, active ▪ can directly cause a disease or can be
▪ relaxed form of heterochromatin a "neutral variant", which means it does
▪ where nucleosomes are evident not cause a disease by itself, but can
cause a disease when paired or if it is
→ Nucleosome
in a close "proximity" with another SNP
▪ composed of 147 DNA base pairs
that is causing a disease, thus called a
→ Histones
linkage disequilibrium
▪ highly conserved low molecular weight
▪ SNPs influence disease susceptibility
proteins
directly
▪ where DNA is wound
• Central dogma of genetics b. Copy number variations (CNV)
1. DNA can be replicated by itself into another DNA ▪ multiple nucleotide polymorphism
2. DNA can also be transcribed into an RNA ▪ long stretches of DNA or different
• importance: after all the splicing and numbers of repeated sequence of DNA
removal of the non-protein coding parts of
the RNA, this RNA is translated into amino Other forms of non-coding DNA:
acids which then forms the protein, in the → Promoters, enhancers, silencers, and binding sites
form of either the cytoskeleton, enzyme, ▪ transcription factors
etc.
→ Telomeres and centromeres
• Genes ▪ special structural regions of DNA
→ contain ~3.2 billion DNA base pairs
▪ 1.5% (20,000): protein-coding genes
▪ 98.5%: for gene regulation
• Gene regulation
1. Non-protein-coding DNA sequences
→ Major classes:
a. Special structural regions of DNA:
▪ Centromeres: chromosome
“tethers”; holds two chromatid
strands together
▪ Telomeres: chromosome ends;
responsible for aging
b. Mobile genetic elements
▪ i.e., transposons: jumping genes
▪ one third of the genome is
composed of such "jumping
genes" that are implicated in the
gene regulation and chromatin
organization

2. Non-protein-coding RNA sequences

→ Varieties:
GENPATH TRANS 1.01 | Trans Team: Alinoor, Alvarez, Banuag, H. Rasul, Macaayan, Macabangon, Nadera, Yap | Editor: Alcuitas, Alvarez, Bernadez 1 of 4
 B. Noncoding RNA o for example: in the laboratory we
• Non-coding RNA want to produce a rat which does
a. Micro-RNA (miRNA) not have any hair. so we use
▪ composed of 21 to 30 nucleotides Knockdown technology to remove
▪ function to modulate the translation of the RNA that is translated into hair
target mRNAs into their corresponding protein.
proteins → Gene silencing
▪ involved in post-transcriptional ▪ functions primarily to modulate the
silencing translation of target mRNAs
o produces a primary mRNA ▪ responsible for posttranscriptional silencing
o fundamental and well- conserved of gene expression
mechanism of gene regulation ▪ binds to a complex called (RISC)
o RNA-induced silencing complex o acts like a transcription factor or
(RISC) polymerase
o subsequent base pairing between o involved in silencing
the miRNA strand and its target ▪ may occur in two ways:
mRNA directs RISC to either 1. Translational repression
induce mRNA cleavage or repress ▪ RISC complex binds with miRNA and
its translation blocks translation
▪ Small interfering RNAs (siRNAs) 2. Complex binds to RNA and causes
o short RNA sequences that can be cleavage
introduced into cells ▪ breaks the RNA that is supposed to be
o synthetically produced miRNA translated into protein
o serves as substrates for Dicer and ▪ the process is called post-
interacts with the RISC complex transcriptional silencing or gene
o “Knockdown technology”: silencing
synthetic siRNAs targeted against
specific mRNA species have The concept of miRNA is illustrated in this diagram:
become useful laboratory tools to
study gene function → it is worth noting that there are two genes in this
diagram: the target gene and the miRNA gene
▪ transcription of target genes results in the
production of target mRNA, which is then
translated into protein
▪ miRNA genes are transcribed to produce
primary miRNA (pri-miRNA) and then pre-
miRNA
▪ through dicer enzyme, pre-miRNA is
spliced to generate a mature miRNA
▪ miRNA binds to RISC to block translation
or break RNAs that are translated into
proteins -> post-transcriptional silencing or
gene silencing

b. Long non-coding RNA (lncRNA)

▪ recent studies have further identified an
untapped universe of lncRNAs which
exceed coding mRNAs by 10-fold to 20-
fold
▪ it is long because it is composed of not just
one single nucleotide but a lot and linked
together.

→ Functions of lncRNA:

a) Gene Activation
▪ enhancers as sites of IncRNA
▪ lncRNAs bind to ribonucleoprotein transcription
complexes similar to RISC complex which is
involved in transcription and gene activation
▪ when genes are activated, transcription and
translation will proceed.
GENPATH TRANS 1.01 | Trans Team: Alinoor, Alvarez, Banuag, H. Rasul, Macaayan, Macabangon, Nadera, Yap | Editor: Alcuitas, Alvarez, Bernadez 2 of 4
 b) Gene Suppression
▪ lncRNAs can preemptively bind transcription
factors to inhibit transcription
▪ transcription and translation will not proceed
▪ i.e., X-inactive specific transcript (XIST)
c) Promote chromatin modification
▪ binding of the lncRNAs and transcription
complex causes changes in the histones
called methylation and acetylation (Picture: There are 9 units of histones, H1 being the
d) Assembly of protein complexes linker histone)
▪ lncRNAs can act as scaffolds to stabilize
secondary or tertiary structures and multi- → there are approximately two turns of DNA
subunit complexes that influence chromatin around a histone (1.8 turns, about 150 base
architecture or gene activity pairs)
▪ this complex becomes an attachment of other → when a DNA from one cell is stretched, it can
transcription factors causing changes in the be as long as 2 meters but if it's clumped in a
DNA. nucleus, it can be as small as 8 micrometers.
→ the histone units are positively charged, thus
allowing compaction of the negatively charged
DNA

Two Types of Chromatin:

1. Heterochromatin
▪ inactive form
▪ the methyl group in the heterochromatin
maintains its inactive form. It cannot be
translated since it is in the clumped form.
2. Euchromatin
▪ active form
▪ it has acetyl groups which promote
relaxation of the DNA in euchromatin.
▪ because only euchromatin permits gene
expression and thereby dictates cellular
identity and activity, there are mechanisms
that regulate the state of chromatin:

a. Histones and histone modifying factors

▪ Chromatin remodeling complexes
o can reposition nucleosomes on
DNA, thus exposing (or
obscuring) gene regulatory
elements, such as promoters
▪ “Chromatin writer” complexes/
Writers/ Marks
o carry out more than 70 different
C. Epigenetics
histone modifications denoted as
marks
• IMPORTANCE OF EPIGENETICS IN REAL LIFE
o covalent alterations include
→ when epigenetics is dysregulated, there is methylation, acetylation, or
disturbance in the epigenome, and this causes phosphorylation of specific
diseases amino acid residues on the
→ it has a central role in cancer and in other diseases histones
o marks are reversible through the
activity of “chromatin erasers”
o “chromatin readers” bind
marked histones, thereby
regulating gene expression
b. Histone methylation
▪ lysine and arginine can be
methylated

GENPATH TRANS 1.01 | Trans Team: Alinoor, Alvarez, Banuag, H. Rasul, Macaayan, Macabangon, Nadera, Yap | Editor: Alcuitas, Alvarez, Bernadez 3 of 4
 ▪ methylation of lysine residues may
be associated with transcriptional
activation or repression
▪ SILENCING
c. Histone acetylation
▪ lysine residues are acetylated by
histone acetyl transferases (HAT),
which opens up chromatin thereby
increasing transcription
▪ can be reversed by histone
deacetylases (HDAC), leading to
chromatin condensation
▪ ACTIVATION
d. Histone phosphorylation
▪ serine residues can be
phosphorylated, which can cause the
DNA to either be opened up for
transcription or condensed to
become inactive
▪ ACTIVATION OR INACTIVATION
e. DNA methylation
▪ high levels of DNA methylation in
gene regulatory elements can cause II. GENE EDITING
silencing
▪ tightly regulated by CONCEPT: Gene editing is basically on how a certain
methyltransferases, demethylating bacteria fights its enemy.
enzymes, and methylated-DNA-
binding proteins • Self vs Enemy
▪ SILENCING → Bacteria vs Phages
f. Chromatin organizing factors → Bacteria takes in phage’s DNA from previous
▪ believed to bind to non-coding attack & inserts it into its own DNA as Clustered
regions and control long-range Interspersed Short Palindromic Repeats
looping of DNA, which is important (CRISPR) -> creates MEMORY
in regulating spatial relationships → CRISPR then is being transcribed into a guide
between gene enhancers and RNA (gRNA)
promoters → gRNA becomes bound into a protein called
Cas9 nuclease, cleaves the current phage’s
DNA -> leads to mutation
→ this complex is the one responsible in fighting
the next virus that it encounters
→ Mutations in Enemy’s DNA:
▪ Non-Homologous End-joining (NHE)
mechanism
o error prone: DNA with random
• Both genetics (DNA) and epigenetics (environment), mutation
play an important role in the development of a person ▪ Homologous DNA recombination
▪ for example, a person might have good genes, o precise changes: DNA with specific
but these genes might be affected by vices, mutations
medications, traumas, and stress. This can lead
to deterioration of the DNA. This also explains You might wonder how this concept in gene editing
why monozygotic twins differ from each other among bacteria becomes applicable in humans, imagine
when they grow up, because of environmental the RNA of the certain cell without disease (refer to the
differences. picture), and a protein where it binds to (gRNA), and the
DNA of a cell, a certain segment which contains DNA that
will be coded for cancer or diabetes. The portion (double-
stranded DNA break :NHEJ) undergoes mutation,
therefore it doesn’t become a diabetic cell or a cell
transformed into cancer.

GENPATH TRANS 1.01 | Trans Team: Alinoor, Alvarez, Banuag, H. Rasul, Macaayan, Macabangon, Nadera, Yap | Editor: Alcuitas, Alvarez, Bernadez 4 of 4