Salk研究所與加州大學(xué)圣地亞哥分校的研究人員合作，利用高分辨率的3D影像,，捕捉神經(jīng)系統(tǒng)發(fā)育過程中重要的蛋白質(zhì)開關(guān)Scp1之作用,。他們的研究結(jié)果發(fā)表于12月8 日的Molecular Cell中,，為Scp1之小分子抑制劑的設(shè)計(jì)提供了一個(gè)模板。
Scp1 是調(diào)控神經(jīng)細(xì)胞前體發(fā)育成為成熟神經(jīng)元的重要開關(guān),。開啟及抑制這個(gè)開關(guān)會(huì)影響神經(jīng)細(xì)胞的分化時(shí)間,。
分子開關(guān)控制著胚胎干細(xì)胞分化成不同的組織類型。因此操控不同的開關(guān)將可以讓科學(xué)家轉(zhuǎn)換胚胎干細(xì)胞成為特殊的細(xì)胞類型,。
研究人員于去年時(shí)發(fā)現(xiàn),，Scp1可以靜默非神經(jīng)細(xì)胞的神經(jīng)元特殊基因。如果能利用特異性抑制劑,，將有助于研究人員觀察胎兒神經(jīng)系統(tǒng)的發(fā)展,，也可以讓醫(yī)學(xué)界將胚胎干細(xì)胞提升為具有特殊功能的神經(jīng)細(xì)胞，而用于治療上,。
Scp1 屬于一組名為小型羧端磷酸酶(SCPs)的蛋白質(zhì),，這種蛋白質(zhì)幾乎在身體的所有組織中表現(xiàn)。當(dāng)Scp1活化時(shí),，可以防止酵素RNA聚合酶 II在神經(jīng)細(xì)胞基因不該表現(xiàn)的區(qū)域如皮膚,、肌肉和肝臟組織中，讀取和啟動(dòng)這些神經(jīng)細(xì)胞基因,。在神經(jīng)系統(tǒng)中,，Scp1被關(guān)閉，使RNA聚合酶 II可以有效地轉(zhuǎn)錄神經(jīng)細(xì)胞的基因信息,，并且使神經(jīng)系統(tǒng)的干細(xì)胞成熟為特殊的神經(jīng)元,。
激酶可以為RNA聚合酶 II加上一個(gè)小的磷酸鹽基團(tuán)，使RNA聚合酶 II繼續(xù)轉(zhuǎn)錄基因,，而Scp1則會(huì)移除小的磷酸鹽基團(tuán),，使RNA聚合酶 II停工。在這項(xiàng)新研究中,，研究人員分析了Scp1和RNA聚合酶 II,，并且獲得了Scp1如何附著于RNA聚合酶 II尾端重復(fù)的七個(gè)胺基酸殘基的3D影像。
研究人員也發(fā)現(xiàn)到,，這七個(gè)胺基酸中只有三個(gè)胺基酸,，與Scp1去除與RNA聚合酶II結(jié)合的磷酸鹽之能力有關(guān)。
英文原文：
Detailed 3-D image catches a key regulator of neural stem cell differentiation in action
La Jolla, CA —Researchers at the Salk Institute for Biological Studies in collaboration with scientists at the University of California, San Diego (UCSD) took a high resolution “action shot” of a protein switch that plays a crucial role in the development of the nervous system. Their findings, published in the Dec. 8 issue of the journal Molecular Cell, provide a template for the design of small molecule inhibitors to control that switch, a protein called Scp1, at will.
“Scp1 is an important brake that regulates the transition from neuronal precursor to mature neuron,” explains senior author Joseph Noel, Ph.D, a Howard Hughes Medical Institute investigator at Salk. “Loosening the brake with an inhibitor would allow us to influence the timing of neuronal differentiation,” he adds.
A finely tuned network of molecular “on” and “off” switches orchestrates the differentiation of embryonic stem cells into different tissue types. Being able to manipulate individual switches would allow scientists to nudge embryonic stem cells into becoming specific cell types, a plus for both basic research and potential therapies.
“At the moment, the differentiation of stem cells into neurons in a Petri dish is a little bit like a black box and not very efficient,” explains co-author Samuel Pfaff, Ph.D., a professor in the Salk’s Gene Expression Laboratory, who together with co-author Gordon Gill, Ph.D., of the Departments of Medicine and Cellular and Molecular Medicine at UCSD, found that Scp1 silences neuron-specific genes in non-neuronal cells last year. “Having a specific inhibitor would give us a lot of insight into the development of the fetal nervous system and would allow us to chemically push embryonic stem cells to acquire a neuronal fate in an informed way,” adds Pfaff.
Scp1 belongs to group of proteins called small carboxyl-terminal phosphatases (SCPs) that are expressed in almost all tissues of the body. When active, Scp1 prevents the enzyme RNA polymerase II from reading and switching on neuronal genes in tissues where they shouldn’t be expressed, such as skin, muscle and liver. In the nervous system Scp1 is switched off, enabling RNA polymerase II to efficiently transcribe information encoded by neuronal genes and driving the maturation of neural stem cells into specialized neurons.
“Scp1 is an interesting twist on how genes can be regulated during development,” says Pfaff. “In the past there has been a lot of emphasis on chromatin modifications and physical access to genes, but Scp1 regulates the activity of the enzyme that transcribes genes directly,” he adds.
Scp1 is not the only protein that directly influences the activity of RNA polymerase II. A constantly fluctuating brigade of enzymatic foot soldiers regulates RNA polymerase II's activity by chemically modifying the long cord-like tail that hangs from its globular structure, like a chain on a light fixture.
Enzymes called kinases turn the “light” on by adding small phosphate chemical groups - giving RNA polymerase the go-ahead to transcribe genes - while removal of those phosphates by phosphatases like Scp1 turns out the light, effectively stopping RNA polymerase in its tracks.
Noel and postdoctoral fellow Yan Zhang, Ph.D, analyzed the crystal structure of Scp1 and RNA polymerase together and obtained a 3-dimensional image showing how Scp1 hangs onto the seven amino acid residues reiterated in the polymerase tail. “We captured Scp1 bound to a single seven amino-acid long repeat containing specific phosphates,” explains Zhang, the paper’s first author. “It turns out that only three amino acids are important for Scp1's ability to know how to remove phosphates from RNA polymerase.”
She adds that knowing how enzymes like Scp1 precisely recognize that seven amino acid stretch is exactly the kind of "unambiguous information relevant for the design of a chemical inhibitor by a process known as structure-based drug design.”
Capitalizing on information gleaned from their structural studies, Noel’s lab has already started that structure-based program. “We have designed the first generation of inhibitors and now it is a matter of chemically synthesizing them, testing them in test tubes and cells, and imaging them bound to Scp1 in 3D,” says Noel. This will set the stage for the rational fine-tuning of their efficacy using an established process Noel likens to molecular dentistry, because his group is striving to shape inhibitory molecules to fit the groove in Scp1 much the same way that a dentist molds a filling to fit the cavity in a patient’s tooth.
Researchers who contributed to the work include Nicolas Genoud, Ph.D., in the Gene Expression Laboratory at the Salk Institute, Jack E. Dixon, Ph.D., and Youngjun Kim, Ph.D., in the Departments of Pharmacology and Cellular and Molecular Medicine at UCSD, and Jianmin Gao, Ph.D., and Jeffery W. Kelly, Ph.D., at the Skaggs Institute for Chemical Biology of The Scripps Research Institute, La Jolla.
The Salk Institute for Biological Studies in La Jolla, California, is an independent nonprofit organization dedicated to fundamental discoveries in the life sciences, the improvement of human health and the training of future generations of researchers. Jonas Salk, M.D., whose polio vaccine all but eradicated the crippling disease poliomyelitis in 1955, opened the Institute in 1965 with a gift of land from the City of San Diego and the financial support of the March of Dimes.
