Characterization of kinesin-like proteins in silkworm posterior silk gland cells.
Kinesins are microtubule-based motors involved in various intracellular transports. Neurons, flagellated cells, and pigment cells have been traditionally used as model systems to study the cellular functions of kinesins. Here, we report silkworm posterior silkgland (PSG), specialized cells with an extensive endomembrane system for intracellular transport and efficient secretion of fibroin, as a novel model for kinesin study. To investigate kinesin-driven intracellular transport in PSG cells, we cloned five silkworm kinesin-like proteins (KLPs), BmKinesin-1, BmKinesin-6, BmKinesin-7, BmKinesin-13, and BmKinesin-14A. We determined their expression patterns by relative real-time PCR and western blotting. Immunofluorescence microscopy verified their colocalization with microtubules. By combining pull-down assays, LC-MS/MS, and western blotting analysis, we identified many potential cargoes of BmKinesin-1 in PSG, including fibroin-containing granules and exuperantia-associated ribonucleoprotein (RNP) complexes. Moreover, BmKinesin-13 overexpression disrupted the microtubule network in BmN cells, which is consistent with a role of Kinesin-13 in regulating microtubule dynamics in other organisms. On the basis of these results, we concluded that PSG might have advantages in elucidating mechanisms of intracellular transport in secretory tissues and could serve as a potential model for kinesin studies.
Wang
Qiao
Q
The Key Laboratory of Cell Proliferation and Differentiation of Ministry of Education, The State Key Laboratory of Bio-membrane and Membrane Bio-engineering, College of Life Sciences, Peking University, Beijing, China.
Teng
Junlin
J
Shen
Birong
B
Zhang
Wei
W
Guo
Yige
Y
Su
Xiaolei
X
Zhang
Chuanxi
C
Yu
Albert C H
AC
Chen
Jianguo
J
D000818:Animals; D001693:Biological Transport, Active / Q000235:genetics / Q000502:physiology; D012831:Bombyx / Q000737:chemistry* / Q000235:genetics / Q000502:physiology; D005088:Exocrine Glands / Q000737:chemistry* / Q000166:cytology* / Q000502:physiology; D006801:Humans; D019476:Insect Proteins / Q000737:chemistry* / Q000235:genetics / Q000502:physiology*; D016547:Kinesins / Q000737:chemistry* / Q000235:genetics / Q000502:physiology*; D005810:Multigene Family / Q000235:genetics; D010802:Phylogeny; D047011:Silk / Q000096:biosynthesis; D022161:Transport Vesicles / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
D019476:Insect Proteins; D047011:Silk; D016547:Kinesins