ゴ
ビ
ン
ダ
シ
ャ
ル
マ
シャルマ
氏
名
ゴビンダ
所
属
理工学研究科 生命科学専攻
学 位 の 種 類
博士（理学）
学 位 記 番 号
理工博 第 217 号
学位授与の日付
平 28 年 9 月 30 日
課程・論文の別
学位規則第４条第 1 項該当
学位論文題名
The
interaction
of
Lemur
kinase
1A
(LMTK1A)
with
cytoskeletons.
Lemur kinase 1A (LMTK1A)の細胞骨格構造との相互作用（英文）
論文審査委員
主査 教 授 久永 眞市
委員 教 授 門田 明雄
委員 教 授 岡本 龍史
委員 准教授 安藤 香奈絵
【論文の内容の要旨】
Summary
Neurons communicate with each other through neurites, axons and dendrites.
Neurite formation requires coordinated actions of cytoskeletons and membrane
trafficking. Two members of cytoskeletons, microtubules and actin filaments serve as
tracks for intracellular membrane traffic depending on regions of neurites, microtubules
in the shaft and actin filaments at the tip. Membrane vesicles used for membrane supply
to the tip of growing neurites are mainly recycling endosomes, which are controlled by
Rab11, a small GTPase. LMTK1A, a novel Ser/Thr kinase expressed abundantly in
brains, regulates trafficking of Rab11-positive endosomes in neurites. However
LMTK1A does not bind directly with Rab11a, and the mechanism how LMTK1A
regulates endosome transport is a question to be answered. In this thesis I investigated
the interaction of LMTK1A with cytoskeletons, tracks of vesicle transport.
At first, I searched for the substrates or proteins interacting with LMTK1A by
immunoprecipitation from the mouse brain membrane fraction, which was prepared by
differential centrifugation of brain lysed in absence of detergent. Mass spectroscopy
revealed many proteins, particularly those related to cytoskeletons or membrane
trafficking. Although I examined the direct interaction of several candidates with
LMTK1A in cultured cells, however, I could not confirm the binding distinctly,
probably due to the difference in sample from brain and from cell culture. Then, I
studied the interaction of LMTK1A with cytoskeletons, particularly microtubules and
actin filaments using cultured PC-12 and Neuro2A cells. I found that LMTK1A affects
microtubule organization in Neuro2A cells. Expression of kinase negative (kn)
LMTK1A shifted the microtubules from the center of cell to the peripheral region.
Concomitantly, γ-tubulin changed its position from underneath of the nucleus to the
lateral side. However it is not clear how microtubule organization is changed by the
expression of kn LMTK1A, one of the reason could be change of γ-tubulin position
relative to nucleus. When γ-tubulin is under nucleus, the astral arrays of microtubules
emanating form γ-tubulin containing MTOC, can spread unhindered, However when
γ-tubulin is beside nucleus, the microtubules need to go around to nucleus, leading to
less amount of microtubules in the central region of the cell. The position of MTOC is
important for polarization of cell. The change in its position in response to LMTK1A
shows that LMTK1A may have a role in cell differentiation.
Further, I found that LMTK1A regulated the transition of endosomal vesicles
from the end of microtubule to the cortical actin-rich region, which would be required
for supply of membrane components for growth of neurite. While wild type (wt)
LMTK1A inhibited such transition, kn LMTK1A could move over to the cortical actin.
Transition of vesicles from microtubule to actin and changing from microtubule
associated motor to actin based motor is limiting step for the unhindered supply of
components for growth of the neurite. Based on my finding, I would like to propose that
LMTK1A enhances minus end-directed movement of Rab11-positive vesicles by
stimulating dynein-dependent movement and regulates the transition of the tracks from
microtubules to actin filaments by suppressing kinesin-dependent movement.