Microtubule-Targetable Fluorescent Probe: Site-Specific Detection and Super-Resolution Imaging of Ultratrace Tubulin in Microtubules of Living Cancer Cells

Abstract

Tubulins in microtubules have been recognized as potential targets in cancer chemotherapy for several years. However, their detection and imaging in living cells, especially following exposure to anticancer drugs, remains difficult to achieve. This difficulty is due to the very small cross section of microtubules and the very small changes in tubulin concentration involved. Photoswitchable fluorescent probes combined with the "super-resolution" fluorescence imaging technique present an exciting opportunity for site-specific detection and super-resolution imaging of specific microscopic populations, such as tubulin. In this study, a tubulin specific photoswitchable fluorescent probe (Tu-SP), that labels and detects ultratrace levels of tubulin in microtubules of living biosystems, was designed and evaluated. To realize super-resolution fluorescence imaging, the spiropyran derivative (SP), a classic photoswitch, was introduced to Tu-SP as a fluorophore. To detect ultratrace tubulin, Tu-SP employed the tubulin inhibitor, alkaloid colchicine (Tu), as a recognition unit. Tu-SP exhibited nearly nonintrinsic fluorescence before binding to tubulin, even if there were divalent metal ions and 375 nm lasers, respectively. After binding to tubulin, a dramatic increase in fluorescence was detected within milliseconds when irradiated at 375 nm, this increase is a result of the transformation of Tu-SP into a colored merocyanine (Tu-SP-1) with fluorescence. Tu-SP was successfully used for site-specific imaging of tubulin at a resolution of 20 ± 5 nm in microtubules of living cancer cells. More importantly, the probe was suitable for site-specific and quantitative detection of trace tubulin in microtubules of living biological samples.

Discerning the Chemistry in Individual Organelles with Small‐Molecule Fluorescent Probes

“…On the basis of the advantages and disadvantages of the present OTAFPs,several future directions in this field are listed herein that could lead to OTAFPs with unique properties: 1) From ap harmacokinetic point of view,d eveloping ap otent and suitable carrier that facilely drives its guest to the target organelle is favorable for maximizing the applicability and minimizing the burden of developing probes.Such ademand has been realized for mitochondria and lysosomes to certain extent, but not for other organelles.V ery recently the Tang research group rationalized as et of fluoride (F À )p robes aiming at either mitochondria or lysosomes,w ith only the organelletargeting group changed (125 and 126,S cheme 45). [241] Pioneering research has revealed the existence of microtubules [242] and lipid droplets [243] within intact live cells,but these organelles should not only be targeted, their physiology should also be studied. 2) Thes ignal-to-noise ratio is ak ey concern of bio-imaging, with autofluorescence from endogenously abundant cellular molecules (e.g.…”

“…[240] Lipid droplets,onthe other hand, are composed of alipid ester core and ap hospholipid monolayer,w hile hosting proteins and being involved in cell metabolism. [241] Pioneering research has revealed the existence of microtubules [242] and lipid droplets [243] within intact live cells,but Angewandte Chemie Reviews these organelles should not only be targeted, their physiology should also be studied.…”

LG308, a Novel Synthetic Compound with Antimicrotubule Activity in Prostate Cancer Cells, Exerts Effective Antitumor Activity

“…In fact, drugs possessing antimicrotubule activity have been used clinically as anticancer drugs for several years (Zhang et al, 2015), but many obstacles to effective treatment with currently approved agents remain, incorporating significant side effects (Risinger et al, 2009;Novio et al, 2014). According to our research, LG308 has acceptable toxicity.…”

Super-resolution fluorescent materials: an insight into design and bioimaging applications

“…Therefore, Moerner et al modified the conventional photoswitchable (UV light) rhodamine fluorogens to develop a new class of rhodamine analogues 17-26 ( Fig. 28 To investigate new fluorescent materials for PALM microscopy that are capable of photoswitching, Zhu et al 29 developed a pH-sensitive, photoswitchable, and water-soluble polymeric nanosystem (39) to confine the acidic lysosome with enhanced resolution (Fig. Additionally, they introduced various conjugated chromophores on the lactam nitrogen of the fluorophore to lower the energy required for photoactivation without inducing any significant effects on their photophysical properties.…”