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Regulation Of Gene Expression > Lesson 9 > Flashcards

Lesson 9 Flashcards

1
Q

where are adult stem cells located?

A

somatic stem cells - only in defined tissues such as haematopoiesis, myogenesis, osteogenesis. etc

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2
Q

what are the four model lineages of stem cells?

A

in muscles, in the skin, in the nervous system, and in hematopoietic stem cells

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3
Q

what are the two parts of the muscle cell lineage?

A

muscle progenitors and the differentiated cells → muscle progenitor is able to differentiate into different cells

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4
Q

what is the main feature of muscle stem cells?

A

the capacity for self-renewal

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5
Q

what are the two levels of stem cells in the skin?

A

at the embryonic level = embryonic epidermal cells and in adults there are the epidermal stem cells (responsible for skin renewal)

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6
Q

where are epidermal stem cells located?

A

in the profound layer of the derma

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7
Q

describe stem cells in the nervous system:

A

neuroepithelial stem cells that give rise to the differentiated cells

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8
Q

why is it easier to study hematopoietic stem cells as opposed to the other three categories?

A

easiest to harvest → the most well known and studied

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9
Q

name some transcription factors of the muscular system important for the expression of genes in the muscular system (do not occur during development and define ms):

A

MyoD, Myogenin, MYF5, PAX3 and PAX7

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10
Q

what are committed progenitors?

A

cells that are not able to self-renew like HSC but have the possibility to differentiate into more than one lineage

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11
Q

what three cell lines can common myeloid progenitors give rise to?

A

mature granulocytes, megakaryocytes, and erythrocytes

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12
Q

what is each differential stem cell state characterized by?

A

using specific factors (not transcription factors) such as cytokines and growth factors which are able to sustain the differentiation and proliferation of the specific lineages

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13
Q

what specific characteristics can be found in stem cells, but not progenitor cells or differentiated cells?

A

stem cells can durably self-renew, they generate daughter stem cells and they can contribute to the pool of differentiated cells

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14
Q

how can you experimentally tell the difference between hematopoietic stem cells, differentiated cells, and progenitors?

A

the use of antibodies → they all express different surface markers

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15
Q

what two ways are differentiated cells and progenitor cells induced in vivo?

A

through intrinsic and extrinsic factors

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16
Q

what are intrinsic factors?

A

present in the cell → regulate master genes of hematopoietic self-renewal or differentiation (transcription factors and miRNA)

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17
Q

what are some examples of extrinsic factors?

A

growth factors, cytokines, or a complex network of cell-cell interactions

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18
Q

what is the microenvironment of HSC in adult bone marrow called?

A

bone marrow niche

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19
Q

what do HSC serve as in the bone marrow?

A

the stromal component, the vascular component, and the osteoblastic compotent

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20
Q

in the molecular regulation of hematopoiesis, what are the critical transcription factor required for?

A

the formation and function of hematopoietic stem cells

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21
Q

in the molecular regulation of hematopoiesis, what are the other classes of transcription factors necessary for?

A

lineage specific differentiation

22
Q

what percentage of the bone marrow do hematopoietic stem cells make up?

A

1-2%

23
Q

describe a gain of function approach:

A

the expression of a gene is induced at supra physiological level in order to see the effects of the over expression

24
Q

describe a loss of function approach:

A

to abrogate the expression that particular gene

25
Q

why are vectors derived from viruses used in research?

A

a virus’ job is to transfer their host genome into a host cell

26
Q

what are some of the most powerful viral systems we can use for hematopoietic research?

A

Mo-MLV and HIV-1

27
Q

what are vectors derived from Mo-MLV?

A

derived from a family of retroviruses, particularly from the Moloney leukemia virus (virus affecting mice that carries oncogenes and causes leukemia)

28
Q

describe the HIV-1 vector:

A

a human virus belonging to the Lentivirus class which is part of the retroviral family

29
Q

which is more complex, Mo-MLV or HIV-1?

A

HIV-1

30
Q

what are the two components of HIV-1?

A

cis-acting sequences and trans-acting sequences

31
Q

what are trans-acting sequences?

A

sequences required for the formation of the viral particle gag-pol-env

32
Q

what are cis-acting sequences in HIV-1?

A

viral sequences required for the assembly of the viral genome into the viral particles

33
Q

what are the viral particles of the cis-acting sequences?

A

-PBS (primer binding site)
-DIS (genome dimerization signal)
-Ψ (packaging signal)
-PPT (polypurine track, downstream env)
-polyA (poly adenilation sequence, in R)
-U & R (unique and redundant regions)
-SD and SA (splice donor and acceptor, scattered)

34
Q

what is the most important cis-acting sequence signal?

A

Ψ signal → signal in the RNA genome that allow this genome to be included into the viral particle

35
Q

what are trans-acting sequences?

A

gag, pol and env are genes that you can remove from the original genome and insert in another construct ( a plasmid) in order to complement the function

36
Q

the expression of your construct of interest will be driven by what?

A

the strong viral promotor

37
Q

what is the first step in the gain of function approach:

A

clone the sequence

38
Q

in order to test the function of a stem cell, what must you look at?

A

the self-renewal and the differentiation

39
Q

what is the only way to test the self-renewal of stem cells?

A

transfer them into an animal model

40
Q

what is used to to reconstitute the haematopoiesis starting from the transplanted cells and begining with the elimination of the endogenous marrow, in order to prevent/avoid the competition among the transplanted cells and the wt cells of the recipient?

A

chemotherapy or irradiations

41
Q

how can you detect the transplanted cells in the recipient mice?

A

you have different markers and GFP

42
Q

when doing experiments with miRNA what is important to remember?

A

not only the expression matters but the level of expression as well → ex: miRNA expressed at low levels can be okay, but when expressed in higher levels induced leukemia

43
Q

why is the level of expression of miRNA important?

A

important to confer advantages at the hematopoietic stem cells, but if the level is dysregulated the advantages can turn into neoplastic growth

44
Q

where are the three sources of hematopoietic stem cells in humans?

A

core blood, bone marrow, and peripheral blood

45
Q

what is cord blood?

A

in the cord blood we have circulating hematopoietic stem cells, reason why, once the baby is born, cord blood can be conserved. Cord blood is a good source of stem cells even if a unit of cord blood is not sufficient to transplant an adult

46
Q

what is the specific surface marker found in cord blood, bone marrow, and peripheral blood?

A

CD34 - expressed mostly in stem cells

47
Q

why is CD34 useful?

A

you can isolate a population of cells particularly enriched in hematopoietic stem cells

48
Q

what can occur after labeling cells with CD34?

A

you can extract primitive cells from core blood

49
Q

what is the best technique you can use to analyze the expression of a gene once you have already known the gene of interest?

A

qPCR

50
Q

when performing qPCR with the gene of interest, what is the aim?

A

to target CD34 positive cells → detect which are the miRNAs expressed only in the positive fraction

51
Q

what is a human mouse chimera?

A

a mouse that can accept the transplantation of human cells

52
Q

how do human mouse chimera work?

A

this strain of mice is immunodeficient for B, T, and NK murine cells

Regulation Of Gene Expression (28 decks)

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