Image: Not only did researchers at UCSD and UCI visualize a mechanically induced signal traverse human cells, but they also showed that actin filaments and microtubules are involved in the process.

Human cells filmed instantly messaging for first time

Cells tugged in one direction sent biochemical signals in the opposite direction in the form of a signature pattern of fluorescent light

Researchers at UCSD and UC Irvine have captured on video for the first time chemical signals that traverse human cells in response to tiny mechanical jabs, like waves spreading from pebbles tossed into a pond. The scientists released the videos and technical details that explain how the visualization effect was created as part of a paper published in the April 21 issue of Nature.

The researchers working at the UCSD Jacobs School of Engineering's Department of Bioengineering developed a novel molecular "reporter" system, which allowed the dynamic visualization of the activation of an important protein called Src. Peter Yingxiao Wang, lead author of the paper and a post-doctoral researcher in UCSD's Jacobs School of Engineering spent two years designing the reporter molecules to light up selectively only when Src was activated, and not other proteins.

Wang and his co-workers first demonstrated that the novel system was effective in visualizing Src activation in response to a known chemical stimulant, epidermal growth factor. Next, they studied the effect of mechanical stimuli on Src activation. Using technology developed at the Beckman Laser Institute at UC Irvine by its founding director Michael Berns, Wang and Elliott Botvinick, a postdoctoral researcher at UCSD Department of Bioengineering and the Beckman Laser Institute at UC Irvine, attached small, sticky beads to cells and gently tugged the beads to and fro with laser power acting as invisible "tweezers." As the laser tweezers moved the beads in one direction, a video camera attached to a specially equipped microscope recorded the dynamic movement of biochemical signals in the opposite direction in the form of a signature pattern of fluorescent light. The fine spatial and temporal resolution was made possible by a technology called fluorescence resonance energy transfer.

"We had no idea what to expect," said Wang. "The first time we saw these incredible waves spreading across the cells I just said 'Whoa, this is amazing.' We expected to see a signal where the tweezers were pulling the beads, but we did not envision such a directional wave propagating away from the beads." Wang worked on this project under the joint advisorship of Shu Chien, a professor of bioengineering and medicine and director of the Whitaker Institute of Biomedical Engineering at UCSD, and Roger Y. Tsien, professor of pharmacology, chemistry, and biochemistry and investigator with the Howard Hughes Medical Institute at UCSD.

Src is one of a large group of enzymes called kinases that attach a phosphate molecule to one or more target proteins in the cell. This phosphorylation reaction typically switches the target protein from inactive to active status. Many diseases can result either when a kinase gene is mutated and can't properly phosphorylate its targets, or when a normal kinase becomes overactive or not sufficiently active. Indeed, Src has been shown to play a key role in cell growth and development, and in the genesis of cancer, atherosclerosis, and many other disease conditions.

"This study amounts to a proof of principle that if we can visualize the activation of one kinase, we can do the same for many others using the same approach," said Chien, the senior author of the paper. Not only are those additional studies expected to reveal temporal and spatial patterns of kinase activation, but Chien also predicted that there will be practical spin offs.

For example, cells usually tightly control the activity of Src, but in certain cancers its activity is abnormality high. "We think that our ability to measure Src activity with this new visualization technique would be useful as a diagnostic test for many cancers," said Chien. The William J. von Liebig Center for Entrepreneurism and Technology Advancement at UCSD's Jacobs School has provided Chien and Wang with funding to commercialize the new visualization technology as a cancer-detection tool.

The researchers showed that actin filaments and microtubules, structural elements that traverse cells like the ribs of an umbrella, could function as conduits for the spread of biochemical signals. Indeed, when Wang disrupted either actin filaments or microtubules in his test cells, the activation signal no longer spread across the cell. These results suggest that the activation of Src traverses these filamentous structures.

Source: University of California - San Diego

    美國加州大學(xué)的圣地亞哥分校（UCSD）及歐文分校的研究人員首次視頻捕獲到響應(yīng)微小機(jī)械尖刺穿過人類細(xì)胞的化學(xué)信號,，它們就像鵝卵石投入池塘中激起的波紋,。他們的研究論文發(fā)表在2005年4月21日的《自然》雜志上,，論文主要作者是UCSD雅各布斯工程學(xué)院的博士后研究員Peter Wang,。

    UCSD雅各布斯工程學(xué)院生物工程系的研人員開發(fā)了一種新的分子“報(bào)道”系統(tǒng),，這種系統(tǒng)能動態(tài)顯示Src蛋白質(zhì)激活情況。Wang花了兩年時(shí)間使報(bào)道分子能夠僅在Src而非其他蛋白質(zhì)被激活時(shí)才發(fā)出指示,。

    Wang及其同事首先證明新系統(tǒng)能有效顯示響應(yīng)一種已知的化學(xué)刺激物――表皮生長因子的Src激活的情況,。然后，他們研究了機(jī)械刺激對Src激活的效果,。采用加州大學(xué)歐文分校貝克曼激光研究所研制出來的技術(shù),，Wang和另一位博士后研究員Elliott Botvinick將一些粘性小珠, 固定在細(xì)胞上，然后利用激光源（無形的“鑷子”）來回拉動這些小珠,。當(dāng)激光鑷子使這些小珠朝一個(gè)方向移動時(shí),，固定在一個(gè)專用顯微鏡上的攝像頭就會拍攝下生化信號朝著相反方向的動態(tài)移動，其移動方式具有熒光的特征,。因此,，采用熒光共振能量轉(zhuǎn)移（FRET）技術(shù)，就有可能實(shí)現(xiàn)高空間分辯率和高瞬時(shí)清晰度,。

    Wang說,，當(dāng)他們第一次看見這些不可思議的波紋穿越細(xì)胞擴(kuò)散開來時(shí)，真是覺得太奇妙了,，他們根本沒想到會看見從小珠發(fā)散出來的定向波紋,。

    Src是一種激酶，它將磷酸鹽分子固定到細(xì)胞內(nèi)的一個(gè)或多個(gè)目標(biāo)蛋白質(zhì)上（稱為“磷酸化”）,，從而激活目標(biāo)蛋白質(zhì),。如果激酶基因變異或不能正常磷酸化其目標(biāo)，或激酶過于活躍,，或激酶不夠活躍時(shí),，都會導(dǎo)致多種疾病。事實(shí)上,，Src在細(xì)胞生長發(fā)育,，以及癌癥、動脈硬化及其他許多疾病的產(chǎn)生中都起到重要作用,。

    論文另一作者Chien說,，這一衡量Src活性的可視化技術(shù)可用于許多癌癥的診斷。細(xì)胞通常嚴(yán)格控制Src的活性,，但是,，在某些癌癥情況下，Src的活性異常高,。Chien和Wang已獲得將這一可視化新技術(shù)作為癌癥診斷工具來商業(yè)化的基金,。

    這些研究人員還發(fā)現(xiàn)，肌纖蛋白絲和微管（橫貫細(xì)胞的結(jié)構(gòu)部件，就好象雨傘的傘骨）可充當(dāng)生化信號的傳播導(dǎo)管,。Wang發(fā)現(xiàn),，當(dāng)他打斷試驗(yàn)細(xì)胞中的肌纖蛋白絲或微管時(shí)，激活信號就不再穿越細(xì)胞擴(kuò)散,。這說明Src激活橫穿了這些纖維結(jié)構(gòu),。