Posted on 05/17/2009 10:57:24 PM PDT by neverdem
miRNA-145 represses Klf-4, Oct4 and Sox2, and is repressed by Oct4
How pluripotency is controlled is one of biology's major puzzles. New research led by Kenneth Kosik of the University of California, Santa Barbara reveals not only a new piece in the puzzle but also how some major pieces fit together.
A set of transcription factors — Klf4, Oct4 and Sox2 — is sufficient to reprogram specialized cells to pluripotency. As pluripotent cells take on a differentiated fate, these transcription factors disappear. Kosik found that one microRNA (miRNA) is able to repress all three of these proteins.
Although these small RNA molecules can silence gene expression in all sorts of cells, their role in pluripotent stem cells is unclear. Kosik screened for miRNAs whose expression increased as embryonic stems cells differentiated.
One of these was miRNA-145, which exists in frogs, fish and mammals and is also enriched in many germline tissues. Computational tools predicted that it would hit mRNA transcripts for Klf4, Oct4 and Sox2.
To make sure that miRNA-145 interacted with the pluripotency proteins, the researchers attached the untranslated regions of Klf4, Oct4 and Sox2 onto the gene for luciferase and found that miRNA-145 repressed luciferase expression in several types of cells, whereas other miRNA constructs did not. Many more experiments, involving mutating the untranslated regions and upregulating and downregulating miRNA-145, all provided evidence that miRNA-145 inhibits these core pluripotency factors.
"[The work] is exciting for many reasons," says Hannele Ruohola-Baker, who studies stem cells and miRNAs at the University of Washington in Seattle. This knowledge could be exploited not only to understand what keeps cells pluripotent or prompts them to differentiate, but also to make cell therapy safer by providing a mechanism to eliminate unwanted stem cells.
Right now, though, she says the research is at an exploratory stage, with researchers just starting to uncover the miRNAs that control stem cell states. "Let-7, miRNA-21 and now miRNA-145 have all been shown to be miRNAs that are not expressed in stem cells and, at varying levels of certainty, have been shown to make cells differentiate," she says. "Who knows, maybe this one [miRNA-145] is the most powerful so far."
And miRNA-145 seems to have a particularly interesting interaction with stem cells' pluripotency machinery. Further study of the genome identified an Oct4 binding site near the part of the genome coding for miRNA-145. Using a luciferase reporter tied to the miRNA-145 region, the researchers found that higher levels of Nanog did not affect the miRNA's expression, but higher levels of Oct4 did. Thus, miRNA-145 represses three powerful pluripotency transcription factors and is itself repressed by one of them — Oct4.
Kosik says that the identification of miRNA-145 and this interesting mechanism opens up many additional questions. "What we need to do now is understand how the double-negative feedback loop is creating a homeostatic state or an altered state. To do that we need a lot more information."
For instance, he says, the number of copies of the various components could help set the balance between the differentiated state and the pluripotent state. Understanding that kind of stoichiometry will be difficult but could be essential to manipulating miRNAs.
There is a precedent for this sort of feedback loop in murine embryonic stem cells, says Isidore Rigoutsos, manager of the bioinformatics and pattern discovery group at the IBM Thomas J. Watson Research Center in Yorktown Heights, New York. He and others have shown that miRNA-296 targets Nanog and that Nanog and Oct4 regulate miRNA-296 (refs 2,3).
Still, Kosik's work is "particularly exciting," Rigoutsos says. "It shows that the details of a feedback loop may change fairly drastically from organism to organism even if one confines oneself to studying specific genes." Understanding such differences could be crucial to manipulating cell states for applications from disease modeling to drug discovery to cell therapy.
Versatile microRNAs, two-faced transcription factors and embryonic stem cells
MicroRNAs boost reprogramming, boot out cMyc
A gene used to reprogram differentiated cells blocks microRNA processing
Efficient ES cell differentiation: the right tweak at the right time
Haifan Lin: peeling back layers of stem cell control
MicroRNAs help embryonic stem cells grow fast
MicroRNA reins in tumor-initiating cells
Repressing microRNAs for pluripotency
Who woulda thought?
stem cell ping
Sounds like my nephew’s PhD thesis. Couldn’t understand that either.
Can't we all work together to try to reduce the number of unwanted stem cells?
The way I’m reading this is that we may have a new path to program stem cells without being dependent on umbilical cords and such. If you have more med schooling than me I’m sure the audience at large would appreciate further illumination.
I heard nothing about stem cells in med school. Between "cancer" and regenerative medicine, I just find the biology fascinating. I have major problems with human embryonic stem cell research. I could not do it for ethical reasons, but others have no such moral objections.
Knowledge could be gained when others engage in human embryonic stem cell research. Any knowledge gained was already bought by a human sacrifice. It will be published as sure as the day is long. I don't have a problem using that knowledge for therapeutic cloning with induced pluripotent stem cells harvested from the same patients with traumatic amputations or severed spinal cords.
They found a way to reprogram somatic cells. Using viral vectors originally to incorporate transcription factors, but lately avoiding viral vectors, they transformed differentiated somatic cells to a state that appears to be equivalent to human embryonic stem cells. They are called induced pluripotent stem cells. I posted articles about them using the keyword ipsc.
A moose bit my sister once.
*****************************EXCERPT**********************
Pluripotency in the broad sense refers to "having more than one potential outcome." In biological systems, this can refer either to cells or to biological compounds. From the Latin pluri=many, potent=power, capacity. A pluripotent cell can create all cell types except for extra embryonic tissue, unlike a totipotent cell, (tot=all), which can produce every cell type including extra embryonic tissue.
Thanks for the link.
Cheers!
BTTT
Mmm, food for thought. The media made it seem as if the stem cell problem with ethics was beaten. Personally I’m freaked out by anything using viruses. Viruses are the most alien form of life (or death?) on our planet. No one really knows why they are and why they continue because they’re not truly alive as we understand life. Yet there they are.
Sorry about the med school comment...closest I’ve come to a lie here. I never snagged the financing but do try to stay read up. Thanks for keeping we morons informed. Somatic - I have been really sleepy of late.
Bad joking aside (and apologizing for the abundance of smartasses who cropped up here) I’d like to know more about pluripotent cells. Do you think that line of research will yield results? There are so many people in need of a cure.
If they can do it without causing a teratoma, yes. They may have to figure out all of the genetics and epigenetics of embryogenesis first.
Somites are pre-definite tissue blocks - but I still get sleepy thinking about them. Such a compicated subject for a Jethro like me. I guess what I’m asking is if we’re any closer to directing blastulation or able to “write over” the varius genesis processes. Pluripotent cells are the point where matter becomes life (do please correct me if I’m wrong). I suspect some things will remain in the realm of God - who created all that in the first place. That we scratch the surface to find a whole new world, every time, says a lot. Says we are perhaps being mocked by the writer.
This is a very disturbing field for mankind to be fishing in but, as we’re all stuck on this one very vulnerable rock, I can’t say we’re wrong trying to insure the perpetuation of our species in every way that doesn’t harm others. Probabilities aside nothing tells us life exists anywhere else. We’re racing against the inevitable asteroid. Meanwhile we have all these genetic and neurological tragedies to care for.
Is it more sinful to fix them or not, if we can? There’s a good question. The Writer’s being silent on this issue of tissue.
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