Seminario biología molecular, tercer semestre, Laura Duque Echeverri, UPB

1. Laura Duque Echeverri | Tercer semestre | Facultad medicina | UPB

2. INTRODUCTION | HDV2
HEPATITIS DELTA VIRUS (HDV) is a satellite of Hepatitis B
virus with a single-stranded circular RNA genome codes for
only one protein, the HDV antigen (HDAg), devoid of any
RNA polymerase activity
HEPATITIS D is an infection that causes the liver to
become inflamed. This swelling can impair liver
function and cause long-term liver problems, including
liver scarring and cancer. It can only infect people who
are already infected with hepatitis B.

3. INTRODUCTION | RNA REPLICATION
3

4. Observe how Hepatitis Delta Virus
histone mimicry drives the
recruitment of chromatin remodelers
for viral RNA replication
GENERAL OBJECTIVE

5. INMUNOFLUORESENCE
FUNDAMENT
Uses antibodies conjugated to
fluorescent dyes to evaluate
cells in suspension, cultured
cells, tissue, for the detection of
specific proteins.
UTILITY
Localize Wt S-HDAg proteins in
the nucleus, using Huh7 cells
using an HDAg antibody and
nuclei stained using DAPI
MATERIAL AND METHODS
5

6. MATERIAL AND METHODS
6
PCR
FUNDAMENT
In vitro enzymatic reaction that
amplifies millions of times a specific
DNA sequence or indirectly through
RNA. Known as RT-PCR by reverse
transcriptase.
UTILITY
To identify which isoform of SNF2L
is/are expressed in hepatocytes,
reverse transcriptase-PCR (RT-PCR)
on total RNA extracted from PHHs,
Huh7, and HepaRG.

7. MATERIAL AND METHODS7
WESTERN BLOTH
FUNDAMENT
Analytical technique used to
detect a specific protein based
on its molecular weight through
the use of antibodies
UTILITY
Analysis of subcellular fractions
to see if ST-S-HDAg, such as
wild-type (wt) S-HDAg,
accumulates in nuclear fractions

8. MATERIAL AND METHODS
8
NORTHEN BLOTH
FUNDAMENT
Identification of specific RNA
sequences, does nos uses restrictive
enzymes due to the small size of
RNA molecules compared to the
DNA.
UTILITY
To assess the effect of R75A
substitution in S-HDAg on HDV RNA
replication

9. Input and HDAg lanes represent
10% and 20% volume compared
with the other lanes, respectively
a
RESULTS PCR products demonstrating that both
SNF2L and SNF2L (ex13) isoforms
were observed in each cell line
b

10. RESULTS
The 4 eluate, shows that all the
proteins are present in exception of
the original histone chain, represents
the level of infection, that the virus
proteins are present using both IB.

11. RESULTS

12. RESULTS
PHHS Infection
During long and short
exposition to the virus, the
Wt RNA presents the major
concentration of the HDAags
inside the infected cells .

13. DISCUSSION
13
AUTHOR DISCUSSION AGREE OR NOT
Huang, W.-H., Mai, R.-T. & Lee, Y.-H.
W
The acetylation of the S-HDAg protein
plays a crucial role in the synthesis of
HDV RNA by cellular Pol II
Schaefer, U., Ho, J. S. Y., Prinjha, R.
K. & Tarakhovsky, A.
Histone mimicry is used by certain
viruses to subvert normal host
defenses
Oppikofer, M. et al.
S-HDAg acetylation motif K72acXXR
mimics the histone KacXXR SLiM and is
read by the BRD of the BAZ2B protein, the
regulatory subunit of the BRF chromatin
remodeling com- plexes18.

14. CONCLUTION
Histone mimicry
The HDV uses a mimicry to the Histone 3
located in the human hepatocytes to
guarantee the RNA replication and to assure
the persitant of the HDV infection, using an
active Polymerase II
Effects of BAZ2B during HDV replication
BAZ2B enhances HDV replication and its BRD
interacts with S-HDAg promoting the disease,
reasured during the whole experiment by the
methods and the innoculation of the healthy
hepatocytes with the mutant genes for the
HDV showing the interaction of the BRD within
the cells original histone strand.
14

15. HEPATITIS DELTA
VIRUS (HDV)
DISEASE
Causes
HEPATITIS D
The liver
Affects
Problems
Cirhosis Cancer Jaundice
Dependsof
Hepatitis B virus
Viral agent
Is a
Requires
Single
stranded
Circular RNA
Codes 1 protein
HDV antigen
Host Cells
Entry
RNA Replication
Hepatocites
S-HDV ag
Through
Using
BAZ2B
Binds
Recognizes
histones
Similiar motif
Using
Histone 3
Foundin
Polymerase II
Activates

16. BIBLIOGRAPHY
▹ https://www.who.int/news-room/fact-
sheets/detail/hepatitis-d
▹ https://www.sciencedirect.com/topics/immunology-and-
microbiology/immunofluorescence
▹ https://www.gesundheitsindustrie-
bw.de/en/article/dossier/dna-and-rna-replication