Spindle fibers are thin, thread-like structures made up of microtubules that are involved in the separation of chromosomes during cell division.

• They are one of the major components of the cell division machinery, formed by the reorganization of microtubules of the cytoskeleton during mitosis and meiosis.
• The microtubules are hollow protein tubes composed of α-tubulin and β-tubulin.
• These fibers are dynamic structures that undergo continuous assembly and disassembly during cell division.
• The spindle fibers are generally arranged in a bipolar manner, extending from the two opposite poles towards the middle region of the dividing cell. Some fibers are attached to the chromosomes at specific regions called kinetochores.
• Other microtubules extend towards the opposite pole and overlap with each other, while some spread out towards the cell periphery.
• The complete arrangement forms a characteristic spindle-shaped structure with a broader middle region and two narrow ends.
• The individual spindle fibers may consist of a single microtubule or a bundle of several microtubules. However, the complete mitotic spindle (spindle apparatus) consists of the organized spindle microtubules, their associated proteins and the structures responsible for their organization at the spindle poles.
• The complete structure is also historically referred to as the “achromatic spindle”, a term used because the spindle does not stain strongly like the chromosomes.
• During meiosis, the corresponding structure is known as the meiotic spindle.

Structure of Spindle Fibers

Spindle fibers are made up of microtubules, which are the components of the cytoskeleton. Their structure and arrangement within the dividing cell are described as follows-

• Microtubule composition- The major protein forming the spindle microtubules is tubulin. It is made up of two globular protein subunits, α-tubulin and β-tubulin, which combine together forming a heterodimer. These tubulin dimers are joined in a head-to-tail arrangement to produce long chains known as “protofilaments”.
• Tubular structure- A microtubule is a hollow cylindrical structure, generally consisting of 13 protofilaments arranged parallel to each other around a hollow central region. It has a diameter of about 25 nm. The microtubules may be present individually or grouped together in bundles within the spindle.
• Polarity and dynamic nature- There are two structurally different ends of the microtubules, the plus (+) end and minus (-) end. The plus end is comparatively more dynamic than the minus end. During cell division, the tubulin subunits are continuously added or removed from the microtubules, which results in their growth and shortening. This property is referred to as “dynamic instability”.
• Bipolar arrangement- In animal cells, the spindle microtubules are arranged around two opposite poles forming a spindle-shaped structure, which is broader at the equatorial region and gradually becomes narrower towards both ends. The spindle in plant cells, on the other hand, is generally more barrel-shaped.
• Kinetochore fibers (K-fibers)- These are bundles of parallel microtubules extending from the spindle poles towards the chromosomes. The microtubules are attached to specific protein structures present at the centromeric region of chromosomes, known as “kinetochores”. Their plus ends remain associated with the kinetochores. The bundles extend towards their corresponding spindle poles.
• Interpolar microtubules- From the two opposite spindle poles, the interpolar microtubules extend towards the middle region of the dividing cell, where they overlap with each other in an antiparallel arrangement. They are not directly attached to chromosomes. Some of these overlapping microtubules also form bundles which are laterally associated with the kinetochore fibers.
• Astral microtubules- In most animal cells, the microtubules extend radially from the centrosomes towards the cell cortex, forming a star-shaped arrangement around the spindle poles known as an “aster”.
• Organization of spindle poles- Centrosomes are the major microtubule-organizing centers (MTOCs) in animal cells. They are responsible for the organization of spindle microtubules. The minus ends of many spindle microtubules are directed towards these poles. Higher plant cells, however, lack centrosomes and organize their spindle microtubules through other microtubule-nucleating and organizing mechanisms.
• Associated proteins- Besides the microtubules, different microtubule-associated proteins (MAPs) and motor proteins are present along the spindle fibers. Motor proteins like kinesins and dyneins are involved in the organization, cross-linking and movement of microtubules. These proteins, along with the spindle microtubules, are components of the complete spindle apparatus.

Types of Spindle Fibers

The spindle microtubules are classified into three major types based on their arrangement, position and attachment within the mitotic spindle of animal cells. They are as follows-

1. Kinetochore Fibers (K-fibers)

Kinetochore fibers are bundles of microtubules that attach to the kinetochores of chromosomes. These microtubules extend between the chromosomes and the spindle poles. They are also called “K-fibers”.

Each kinetochore is attached to a bundle containing several microtubules, with their plus ends associated with the kinetochore and the opposite ends extending towards the corresponding spindle pole. During metaphase, the sister kinetochores are attached to microtubules extending from opposite poles. The chromosomes are held near the equatorial region of the spindle.

In anaphase, the kinetochore fibers undergo shortening by the removal of tubulin subunits from the microtubules. The separated sister chromatids move towards their respective poles.

2. Polar or Interpolar Fibers

These are non-kinetochore microtubules that extend towards the middle region of the spindle. They do not attach to the chromosomes.

The polar microtubules arising from the opposite spindle poles enter the central region, where some of them overlap with one another in an antiparallel arrangement. These overlapping microtubules are referred to as “interpolar microtubules” or “overlap microtubules”.

During anaphase, the overlapping microtubules slide past each other with the help of motor proteins, moving the two spindle poles apart. Some interpolar microtubules also form bundles which are laterally associated with the kinetochore fibers. These are known as “bridging fibers”.

3. Astral Fibers

In most animal cells, the microtubules extending radially from the centrosomes towards the cell periphery are referred to as astral microtubules. They form a star-shaped arrangement around each spindle pole called an “aster”.

Unlike the kinetochore fibers, they are not directly attached to the chromosomes. The astral microtubules interact with the cell cortex, and with the help of associated motor proteins, they are involved in the positioning and orientation of the spindle within the dividing cell.

These microtubules also participate in generating the pulling forces that help in the separation of spindle poles.

Formation of Spindle Fibers

Spindle fibers are formed by the assembly and reorganization of the cytoskeletal microtubules during cell division. In animal cells, the major sites for their formation are the centrosomes, which act as microtubule-organizing centers (MTOCs).

The formation of spindle fibers involves the following steps-

1. Duplication and Separation of Centrosomes

Before the beginning of mitosis, the centrosome is duplicated during interphase, forming two centrosomes. These centrosomes start moving apart in prophase and gradually occupy the opposite sides of the nucleus.

During this stage, the existing interphase microtubules are disassembled. New microtubules begin to grow around each centrosome.

2. Formation of Spindle Microtubules

The centrosome consists of two centrioles surrounded by a protein-rich region called pericentriolar material (PCM). It contains γ-tubulin ring complexes (γ-TuRC), which provide the sites for microtubule nucleation.

Here, the free α-tubulin and β-tubulin subunits combine to form tubulin dimers. These dimers are assembled into new microtubules, with their minus ends associated with the centrosomes and the plus ends growing outwards by adding more tubulin subunits.

The microtubules undergo continuous growth and shortening. Tubulin subunits are added and removed during this process.

Apart from the centrosomes, another site of microtubule formation is around the chromosomes. After the breakdown of nuclear envelope, the chromatin-associated Ran-GTP system promotes the formation of microtubules in this region.

Additional microtubules are also formed from the already existing spindle microtubules by a protein complex called augmin.

3. Organization of the Bipolar Spindle

The microtubules growing from the two centrosomes extend towards the middle region of the cell. Some of these microtubules come in contact and overlap with each other in an antiparallel arrangement.

The motor protein kinesin-5 binds to these overlapping microtubules and slides them past one another. This movement pushes the two centrosomes apart. Other motor proteins are involved in collecting and focusing the microtubules towards the spindle poles.

Following the breakdown of the nuclear envelope, the growing microtubules can reach the chromosomes. Some of these microtubules are captured by the kinetochores, forming kinetochore microtubules.

The overlapping microtubules form the interpolar arrangement. Astral microtubules extend out from the centrosomes towards the cell periphery.

Formation of Spindle Fibers in Plant Cells

The higher plant cells lack centrosomes. Here, the spindle microtubules are mainly formed around the nuclear envelope without the involvement of centrosomes.

During prophase, the microtubules accumulate around the nucleus and gradually become arranged into two groups towards its opposite sides. In many flowering plants, these structures are referred to as “polar caps”. The γ-tubulin is involved in the nucleation of microtubules at these regions.

After the nuclear envelope breaks down, the microtubules are further arranged into a bipolar spindle. Some microtubules attach to the kinetochores of chromosomes forming the kinetochore fibers. Motor proteins and microtubule-associated proteins are involved in the organization of spindle microtubules and their polar regions.

Spindle Fibers During Mitosis

The spindle fibers undergo continuous changes in their arrangement and length throughout mitosis. They are formed during prophase, attach to the chromosomes in prometaphase and undergo further changes during chromosome separation.

The different stages of spindle fiber formation and their activities during mitosis are as follows-

1. Prophase

In animal cells, the two centrosomes, which have already been duplicated during interphase, begin moving towards the opposite sides of the nucleus. The long cytoplasmic microtubules of the interphase cell are replaced by numerous short and highly dynamic microtubules.

Around each centrosome, a star-shaped arrangement of microtubules (aster) is formed. The microtubules growing from the two centrosomes come in contact with each other. Motor proteins are involved in sliding the overlapping microtubules, which helps in moving the centrosomes apart and formation of the two spindle poles.

2. Prometaphase

The nuclear envelope breaks down during prometaphase. Now, the spindle microtubules can reach the chromosomes.

Some of these microtubules attach to the protein structures called “kinetochores” present at the centromeric region of chromosomes. This attachment occurs by a process known as “search and capture”, in which the growing microtubules come in contact with the kinetochores and become attached to them.

The attached microtubules form kinetochore fibers (K-fibers). In the meantime, the two sister kinetochores become attached to microtubules extending from opposite spindle poles. Other microtubules overlap in the middle region of the spindle or extend towards the cell periphery.

3. Metaphase

The complete bipolar spindle is now formed, and the chromosomes are arranged at the equatorial region of the cell, known as the “metaphase plate”.

Each sister kinetochore is attached to the microtubules from the opposite spindle pole. The spindle microtubules remain dynamic, undergoing continuous addition and removal of tubulin subunits. Before the beginning of anaphase, the spindle-assembly checkpoint prevents chromosome separation until proper attachment of the chromosomes is established.

4. Anaphase

During anaphase, the cohesin proteins holding the sister chromatids together are broken down, allowing their separation. The kinetochore fibers begin to shorten by the removal of tubulin subunits, which moves the separated chromosomes towards their respective poles. This movement is referred to as “anaphase A”.

At the same time, the overlapping interpolar microtubules slide past one another with the help of motor proteins, pushing the spindle poles apart. In many animal cells, the astral microtubules also participate in pulling the poles towards the opposite sides of the cell.

The separation of spindle poles is known as “anaphase B”. During this stage, the overlapping microtubules are reorganized into bundles forming the central spindle between the separating chromosomes.

5. Telophase

The separated chromosomes reach the opposite spindle poles and begin to decondense. Their attachment with the kinetochore microtubules is lost, and most of the spindle fibers undergo disassembly.

During this process, the nuclear envelopes are re-formed around the two groups of chromosomes. Some of the central spindle microtubules, however, remain between the daughter nuclei. In animal cells, these microtubules are compressed into a compact structure called the “midbody” during cytokinesis. The remaining microtubules are subsequently disassembled during the final separation of the daughter cells.

Spindle Fibers in Plant and Animal Cells

Spindle fibers are present in both plant and animal cells. They are made up of cytoskeletal microtubules which are used for the separation of chromosomes during cell division. The organization and formation of these microtubules differ in plant and animal cells.

Spindle Organization in Animal Cells

The major sites for the formation of spindle fibers in most animal cells are the centrosomes, also known as microtubule-organizing centers (MTOCs). Each centrosome consists of a pair of centrioles surrounded by a protein-rich region called pericentriolar material (PCM). This material contains γ-tubulin complexes, which provide the sites for the formation of new microtubules.

Before mitosis, the centrosomes undergo duplication. These duplicated centrosomes gradually move towards the opposite sides of the nucleus, where a large number of microtubules begin to grow out from them.

Some of the microtubules coming from the opposite centrosomes overlap with each other in the middle region of the dividing cell. With the help of motor proteins, these microtubules are organized into a bipolar spindle.

Around each centrosome, a star-shaped arrangement of microtubules is also formed. It is referred to as an “aster”. The microtubules forming these structures are called astral microtubules, which extend radially towards the cell periphery and interact with the cell cortex. These interactions are involved in controlling the position and orientation of the spindle within the cell.

The centrosomes occupy the two well-defined spindle poles, while the kinetochore microtubules extend towards the chromosomes.

Spindle Organization in Plant Cells

Unlike animal cells, higher plant cells do not contain conventional centrosomes with centrioles. However, they can form a functional mitotic spindle without these structures. In these cells, the nuclear envelope acts as one of the major sites of microtubule formation during the beginning of mitosis.

During prophase, a large number of microtubules accumulate around the nucleus. These microtubules gradually become organized into two loosely arranged groups towards the opposite sides of the nuclear envelope. In many flowering plant cells, these arrangements are referred to as “polar caps”.

The formation of microtubules in these regions involves γ-tubulin ring complexes (γ-TuRC), which provide the sites for microtubule nucleation.

Apart from this, additional microtubules are produced from the already existing microtubules with the help of a protein complex called augmin. The augmin complex recruits γ-tubulin complexes to the existing microtubules for the formation of new microtubules. Motor proteins are also involved in arranging these microtubules within the spindle.

After the breakdown of the nuclear envelope, the microtubules become arranged around the chromosomes. Some of them attach to the kinetochores, forming the kinetochore fibers.

The mitotic spindle of higher plant cells is generally barrel-shaped with comparatively broad polar regions. Prominent astral microtubules are absent in the typical spindle of flowering plant cells. The spindle microtubules are organized into two functional poles without the presence of centrosomes.

Functions of Spindle Fibers

The following are some of the important functions of spindle fibers during cell division:

• Formation of the bipolar spindle- During prophase, the microtubules are organized to form a bipolar spindle between the two opposite poles of the cell. The overlapping microtubules, along with the motor proteins, help in the formation and maintenance of the spindle structure.
• Attachment of chromosomes- The kinetochore microtubules grow towards the chromosomes and attach to specific protein structures called “kinetochores” present at the centromeres. In mitosis, the sister kinetochores are attached to microtubules extending from opposite spindle poles.
• Alignment of chromosomes- The spindle fibers are involved in bringing the chromosomes towards the equatorial region of the cell. This process is referred to as “chromosome congression”. During prometaphase, the chromosomes move along the spindle microtubules with the help of motor proteins and various pulling and pushing forces. The chromosomes are arranged at the metaphase plate before separation.
• Separation of sister chromatids- During anaphase, the kinetochore fibers help in moving the separated sister chromatids towards opposite poles of the dividing cell. The microtubules shorten by the removal of tubulin subunits, and chromosome movement is also assisted by motor proteins. This movement is known as “anaphase A”.
• Separation of spindle poles- The overlapping microtubules from opposite poles slide past one another with the help of motor proteins, pushing the two spindle poles apart. Astral microtubules can also exert pulling forces on the spindle poles. This process occurs during anaphase B.
• Positioning of the spindle- Astral fibers extend from the centrosomes towards the cell cortex. They are involved in the positioning and orientation of the mitotic spindle within many animal cells.
• Determination of the cell division plane- In animal cells, the spindle microtubules also play a role in determining the position of the cleavage furrow during cytokinesis. Depending on the cell type, either the astral microtubules or the overlapping microtubules of the central spindle help in positioning the contractile ring.
• Chromosome segregation during meiosis- Spindle fibers are also involved in the separation of chromosomes during meiotic cell division. During meiosis I, the spindle microtubules help in moving the homologous chromosomes towards opposite poles while the sister chromatids remain attached. In meiosis II, the sister chromatids are separated and move towards the opposite spindle poles.

What Happens When Spindle Fibers Fail?

The failure of spindle fibers can affect chromosome attachment and separation during cell division. Some of the consequences are as follows-

• Failure of spindle formation- The microtubules may form an abnormal spindle with a single pole (monopolar) or more than two poles (multipolar). Normal separation of chromosomes is affected.
• Defective chromosome attachment- Some kinetochores remain unattached, while both sister kinetochores may get attached to microtubules from the same spindle pole. The chromosomes may fail to reach the metaphase plate. These errors are normally corrected before anaphase.
• Mitotic arrest- When kinetochores remain unattached, the spindle assembly checkpoint (SAC) is activated. It prevents the cell from entering anaphase. Uncorrected attachment defects may result in prolonged mitotic arrest.
• Lagging chromosomes- In merotelic attachment, a single kinetochore becomes attached to microtubules arising from both spindle poles. This defect may escape the spindle assembly checkpoint. During anaphase, the chromosome may remain behind due to opposite pulling forces. Some reach the correct pole, while others undergo abnormal segregation.
• Aneuploidy- Improper chromosome segregation may produce daughter cells with abnormal chromosome numbers. One cell may receive an extra chromosome, whereas the other loses it. This condition is referred to as aneuploidy.
• Formation of micronuclei- The chromosomes left behind during anaphase may form separate small nuclei, known as micronuclei. These chromosomes may undergo abnormal DNA replication, DNA damage and sometimes extensive fragmentation or rearrangement.
• Cell death and mitotic slippage- Prolonged mitotic arrest may result in cell death. Some cells, however, come out of mitosis without completing normal chromosome segregation and cytokinesis. This process is called mitotic slippage. Such cells may become tetraploid, containing twice the normal diploid chromosome complement.
• Chromosomal instability- Repeated errors in chromosome segregation cause continuous changes in chromosome number during successive cell divisions. This condition is known as chromosomal instability (CIN), which is found in many cancer cells.

Spindle Fibers at a Glance

References

1. Alberts, B., Johnson, A., Lewis, J., Raff, M., Roberts, K., & Walter, P. (2002). Molecular biology of the cell (4th ed.). Garland Science. https://www.ncbi.nlm.nih.gov/books/NBK21054/toc/
2. Burgess, A., Rasouli, M., & Rogers, S. (2014). Stressing mitosis to death. Frontiers in Oncology, 4, Article 140. https://doi.org/10.3389/fonc.2014.00140
3. Cimini, D. (2023). Twenty years of merotelic kinetochore attachments: A historical perspective. Chromosome Research, 31(3), Article 18. https://doi.org/10.1007/s10577-023-09727-7
4. Clark, M. A., Douglas, M., & Choi, J. (2018). Biology 2e. OpenStax. https://openstax.org/books/biology-2e/pages/10-2-the-cell-cycle
5. Cooper, G. M. (2000). The cell: A molecular approach (2nd ed.). Sinauer Associates. https://www.ncbi.nlm.nih.gov/books/NBK9839/
6. di Pietro, F., Echard, A., & Morin, X. (2016). Regulation of mitotic spindle orientation: An integrated view. EMBO Reports, 17(8), 1106–1130. https://doi.org/10.15252/embr.201642292
7. Ganem, N. J., & Pellman, D. (2012). Linking abnormal mitosis to the acquisition of DNA damage. Journal of Cell Biology, 199(6), 871–881. https://doi.org/10.1083/jcb.201210040
8. Gatlin, J. C., & Bloom, K. (2010). Microtubule motors in eukaryotic spindle assembly and maintenance. Seminars in Cell & Developmental Biology, 21(3), 248–254. https://doi.org/10.1016/j.semcdb.2010.01.015
9. Joglekar, A. P. (2016). A cell biological perspective on past, present and future investigations of the spindle assembly checkpoint. Biology, 5(4), Article 44. https://doi.org/10.3390/biology5040044
10. Lee, Y.-R. J., & Liu, B. (2019). Microtubule nucleation for the assembly of acentrosomal microtubule arrays in plant cells. New Phytologist, 222(4), 1705–1718. https://doi.org/10.1111/nph.15705
11. Maiato, H., Gomes, A. M., Sousa, F., & Barisic, M. (2017). Mechanisms of chromosome congression during mitosis. Biology, 6(1), Article 13. https://doi.org/10.3390/biology6010013
12. Musacchio, A. (2011). Spindle assembly checkpoint: The third decade. Philosophical Transactions of the Royal Society B: Biological Sciences, 366(1584), 3595–3604. https://doi.org/10.1098/rstb.2011.0072
13. Prosser, S. L., & Pelletier, L. (2017). Mitotic spindle assembly in animal cells: A fine balancing act. Nature Reviews Molecular Cell Biology, 18(3), 187–201. https://doi.org/10.1038/nrm.2016.162
14. Salmon, E. D., Cimini, D., Cameron, L. A., & DeLuca, J. G. (2005). Merotelic kinetochores in mammalian tissue cells. Philosophical Transactions of the Royal Society B: Biological Sciences, 360(1455), 553–568. https://doi.org/10.1098/rstb.2004.1610
15. Tanaka, T. U. (2010). Kinetochore–microtubule interactions: Steps towards bi-orientation. The EMBO Journal, 29(24), 4070–4082. https://doi.org/10.1038/emboj.2010.294
16. Thompson, S. L., & Compton, D. A. (2011). Chromosome missegregation in human cells arises through specific types of kinetochore–microtubule attachment errors. Proceedings of the National Academy of Sciences, 108(44), 17974–17978. https://doi.org/10.1073/pnas.1109720108
17. Tolić, I. M. (2018). Mitotic spindle: Kinetochore fibers hold on tight to interpolar bundles. European Biophysics Journal, 47(3), 191–203. https://doi.org/10.1007/s00249-017-1244-4
18. Valdez, V. A., Neahring, L., Petry, S., & Dumont, S. (2023). Mechanisms underlying spindle assembly and robustness. Nature Reviews Molecular Cell Biology, 24(8), 523–542. https://doi.org/10.1038/s41580-023-00584-0
19. Yamada, M., & Goshima, G. (2017). Mitotic spindle assembly in land plants: Molecules and mechanisms. Biology, 6(1), Article 6. https://doi.org/10.3390/biology6010006