MOLECULAR ORGANISATION OF CENTROMERE AND TELOMERE

Centromere

As earlier described, centromere is a unique region of a chromosome characterized by a constriction, where the two chromatids remain joined together after chromosome replication.
On each of these centromeres, specialized DNA-protein complexes known as kinetochores assemble.
Each chromosome has two kinetochores, one on each sister chromatid, facing in opposite directions.
These kinetochores are the sites for binding of kinetochore microtubules (spindle), which assist in the movement of chromosomes during anaphase.
The DNA sequence representing the centromere contain the information specifying the assembly of kinetochore.
These centromeric DNA sequences have been isolated and characterized in a number of organisms but, unlike the telomeric sequences, have not been found to be conserved.

The kinetochore functions as the “microtubule organizing Centre” (MTOC) of the chromosome. Such centrios also occur at the poles, in the vicinity of centriole.
A single microtubule is sufficient to move a chromosome. but, the number of microtubules attached to a single chromosome .
In yeast there is only one microtubule per kinetochore, but in human, 20 to 40 microtubules are found attached to one kinetochore.
The centromere is surrounded by heterochromatin that can be observed by the C-banding technique. Satellite DNA sequences are located on both the sides of centromeres.
The minimum region of a chromosome necessary for centromeric function is called a CEN fragment or CEN region.
For example – In case of yeast, Each of the 17 chromosomes of yeast contains a different centromeric sequence, each sequence being with in 120 bp in length.
However, all the sequences contain substantial regions of homology, and can be inverted or swapped from one chromosome to another without loss of function.
The yeast centromeric DNA contains three distinguishable sequences.

(i) Conserved element I (CDE I):

It is composed of 9 bp and is located at the left end of the centromere; it shows minor variations.

(ii) Conserved element II (CDE II):

This element is the middle region containing 80-90 bp. A=T rich sequences constitute more than 90% of this region.

(iii) Conserved element III (CDE III):

This element consists of 11 bp and is located at the right end of the centromere (Fig. 8.8). It is a highly conserved sequence.

Point mutations in the CCG sequence of the conserved element III leads to an inactivation of the centromeric function.
Mutations in the other elements, however, cause only a reduction in the centromeric function.
Certain proteins or protein complexes bind to specific regions of the CEN sequences.

Protein CBF-I binds to CDEI element. Proteins CBF-IIIA, CBF-IIIB and CBF-IIIC from a complex (M.W. 2.4 × 105 Daltons) which binds to the CDE-III region.
This protein complex has some motor activity due to which the centromeric region of the chromosome becomes attached to microtubules.
Mitotic chromosome movement is inhibited when mutation occurs in the genes coding for CBF-III proteins.
The yeast centromeric sequences bind to specific proteins which initiate to form a kinetochore (multi-protein complex).
The kinetochore, in turn, binds to the end of a single microtubule. Mammalian centromeres are thought to contain different and much longer DNA sequences.
They form larger kinetochores each of which binds to several microtubules.

Telomere

The sequences at the ends of eukaryotic chromosomes, called telomeres, play critical roles in chromosome replication and maintenance.
Telomeres were initially recognized as distinct structures because broken chromosomes were highly unstable in eukaryotic cells, implying that specific sequences are required at normal chromosomal termini.

In recent years the structure of telomeres in a wide variety of organisms has been studied to demonstrate that telomeres are highly conserved elements throughout the eukaryotes, both in structure and function.
Telomeric DNA has been shown to consist of simple randomly repeated sequences, characterized by clusters of G residues in one strand and C residues in the other.
Another feature is a 3’ overhang (12-16 nucleotides in length) of the G-rich strand. For example, the sequence of telomere repeats in humans and other mammals is AGGGTT, and the telomere repeat in Tetrahymena is GGGGTT.
These sequences are repeated hundreds or thousands of times, thus spanning up to several kilobases, and terminate with an overhang of single-stranded DNA.
Recent results suggest that the repeated sequences of telomere DNA form loops at the ends of chromosomes, thereby protecting the chromosome termini from degradation
Telomeres play a critical role in replication of the ends of linear DNA molecules.
The same repeated sequence is found at the ends of all chromosomes in a species and the same telomere sequence may occur in widely divergent species, such as humans, some acellular slime molds (trypanosome) and the fungi like Neurospora.
At every telomere, as much as 10 kilobase of this repeat sequence may occur.
The telomric DNA is also complexed with non-histone proteins,the complex structure being associated with nuclear lamina, as shown in Oxytricha, a ciliated protozoan.
The telomeric DNA is synthesized under the influence of telomerase.
An enzyme which has been shown to be a ribonucleoprotien, whose RNA component works as a template for synthesis of telomeric DNA repeats and protein component has reverse transcriptase (RT) activity.
Recently (In 1997), proteins have been isolated from telomerase in yeast and a ciliated protozoan (Euplotes) which could be the RT component of telomerase.
These were p123 in Euplotes and Est2 in yeast. In recent years, efforts have also been made to understand, how the telomere length is monitored in telomerase, because uncontrolled activity of telomerase will lead to indefinite elongation of telomeres.
Proteins binding to telomere repeats have been identified, which block the elongation of telomeres.
These include rap1p in budding yeast, Taz1p in fission yeast and TRF in humans.
The significance of these proteins lies in the following two observations made in recent years:
(i) Shortening of telomere tract (repeat sequence) is associated with senescence and aging;
(ii) Controlled elongation of telomere is an essential step in tumour formation and oncogenesis.
However, even in tumour cells, this telomere length does not increase in an uncontrolled manner, and the elongation is regulated.
However, we still do not know, ‘how does the cell measure the telomere length, and how is this information used to regulate the length of telomeres’?

🔹 PART A: CENTROMERE STRUCTURE & FUNCTION

Q1. What is the primary function of the centromere in a chromosome?
A. RNA synthesis
B. DNA replication
✅ C. Attachment site for spindle fibers
D. Protein synthesis

Explanation:
The centromere is where kinetochores form, which bind spindle fibers during cell division.

Q2. Which structure forms at the centromere to assist chromosome movement?
A. Telomerase
B. Centrosome
✅ C. Kinetochore
D. Histone octamer

Explanation:
Kinetochore is a protein complex at the centromere that binds spindle fibers.

Q3. (PYQ – CUET 2022) What is the minimum DNA sequence required for centromeric function in yeast called?
✅ A. CEN fragment
B. Telomere repeat
C. ORI
D. Operator

Explanation:
CEN is the essential centromeric region for kinetochore formation and function.

Q4. Which organism has only one microtubule per kinetochore?
✅ A. Yeast
B. Human
C. Fruit fly
D. Mouse

Explanation:
Unlike humans with 20–40 MTs, yeast has a single microtubule per kinetochore.

Q5. What region flanks the centromere and appears in C-banding technique?
✅ A. Heterochromatin
B. Euchromatin
C. Telomeric DNA
D. Satellite body

Explanation:
C-banding stains heterochromatin, especially near centromeres.

Q6. In yeast centromere, which conserved region is A-T rich and central?
A. CDE I
✅ B. CDE II
C. CDE III
D. ORI

Q7. Which conserved region is 11 bp and highly conserved in yeast centromere?
A. CDE I
B. CDE II
✅ C. CDE III
D. CDE 0

Q8. Point mutation in CCG sequence of CDE III leads to:
✅ A. Inactivation of centromere
B. Enhanced replication
C. Increased gene expression
D. Recombination

Q9. Which protein complex binds to the CDE III region in yeast?
✅ A. CBF-III
B. TBP
C. TATA box
D. Rap1

Q10. Which of these is not part of CBF-III complex?
A. CBF-IIIA
B. CBF-IIIB
✅ C. CBF-IV
D. CBF-IIIC

🔹 PART B: KINETOCHORE & MICROTUBULE ORGANIZATION

Q11. Kinetochores function as:
✅ A. Microtubule organizing centers (MTOCs)
B. DNA ligase centers
C. Gene regulatory hubs
D. mRNA editing sites

Q12. In humans, how many microtubules can attach to one kinetochore?
A. 1
B. 5–10
✅ C. 20–40
D. 100

Q13. What binds chromosomes to the spindle fibers?
A. Telomerase
✅ B. Kinetochore
C. Nucleosome
D. Operator

Q14. CBF-I protein binds to which yeast centromere element?
✅ A. CDE I
B. CDE II
C. CDE III
D. None

Q15. Mutations in CBF-III proteins affect:
A. DNA replication
B. Histone folding
✅ C. Mitotic movement
D. Spliceosome formation

🔹 PART C: TELOMERE STRUCTURE & FUNCTION

Q16. Telomeres are made up of:
✅ A. Repetitive G-rich and C-rich sequences
B. mRNA caps
C. Ribosomal DNA
D. Operon sequences

Q17. (PYQ – NEET 2020) The function of telomere is to:
A. Initiate transcription
✅ B. Prevent chromosome degradation and fusion
C. Replicate proteins
D. Increase recombination

Q18. In humans, the telomeric repeat sequence is:
✅ A. AGGGTT
B. TTAGGG
C. GGGGTT
D. ACGTGA

Q19. Which of the following is a 3’ overhang in telomeres?
A. C-strand
✅ B. G-strand
C. T-strand
D. A-strand

Q20. Telomere DNA ends with a:
A. Hairpin loop
✅ B. Loop with single-stranded overhang
C. Centromere
D. Satellite

Q21. The telomeric DNA is synthesized by:
✅ A. Telomerase
B. Ligase
C. Helicase
D. Topoisomerase

Q22. Telomerase is a:
✅ A. Ribonucleoprotein with reverse transcriptase activity
B. Protein kinase
C. RNA polymerase
D. Ligase

Q23. The protein component of telomerase performs:
✅ A. Reverse transcription
B. DNA replication
C. RNA splicing
D. DNA repair

Q24. In yeast, the telomerase reverse transcriptase component is called:
✅ A. Est2
B. p53
C. RAS
D. p123

Q25. Which protozoan has telomerase protein p123?
✅ A. Euplotes
B. Paramecium
C. Plasmodium
D. Tetrahymena

🔹 PART D: TELOMERE REGULATION & DISEASE CONNECTIONS

Q26. Uncontrolled telomerase activity may lead to:
✅ A. Tumor formation
B. Apoptosis
C. Telomere loss
D. DNA repair defect

Q27. Which protein prevents excessive telomere elongation in budding yeast?
✅ A. Rap1p
B. p53
C. H1
D. TERT

Q28. In fission yeast, which protein blocks telomere elongation?
A. Rap1p
✅ B. Taz1p
C. Est2
D. Ku70

Q29. TRF proteins regulate telomere length in:
✅ A. Humans
B. Yeast
C. Tetrahymena
D. E. coli

Q30. What is the telomerase RNA template used for?
✅ A. Adding repeats to telomeres
B. mRNA splicing
C. rRNA synthesis
D. Histone translation

🔹 PART E: TELOMERE LENGTH & AGING

Q31. Shortening of telomeres is associated with:
✅ A. Cellular aging
B. Rapid replication
C. Transcription
D. Ribosome synthesis

Q32. (PYQ – UPSC 2021) Controlled elongation of telomeres is required for:
A. Stem cell quiescence
B. Senescence only
✅ C. Cancer cell immortality
D. DNA repair

Q33. In humans, how long can telomeric repeats span?
✅ A. Up to 10 kb
B. 500 bp
C. 50 kb
D. 100 bp

Q34. Telomere binding proteins regulate length by:
A. DNA methylation
✅ B. Blocking telomerase access
C. Histone binding
D. Transcription activation

Q35. Why are broken chromosomes unstable in eukaryotes?
✅ A. They lack telomeres
B. No centromere
C. Poor RNA structure
D. Lack of G-C base pairing

🔹 PART F: ADDITIONAL CONCEPTUAL MCQs

Q36. Which of the following is a ribonucleoprotein enzyme?
✅ A. Telomerase
B. DNA polymerase
C. Ligase
D. Primase

Q37. Telomeres and centromeres both:
✅ A. Contain non-coding repetitive DNA
B. Encode mRNA
C. Act as operons
D. Are part of ribosomes

Q38. Which chromosomal element helps in preventing fusion between chromosomes?
A. Centromere
✅ B. Telomere
C. Histone
D. Kinetochore

Q39. Which of the following plays a role in spindle fiber attachment?
✅ A. Kinetochore
B. Telomerase
C. H1 histone
D. Rap1

Q40. The loop structure at the end of telomeres helps in:
✅ A. Protecting chromosome ends
B. Forming ribosomes
C. Enhancing RNA transcription
D. Building centrosomes

🔹 PART G: MIXED PYQs + ADVANCED

Q41. Which base is enriched in telomeric sequences?
✅ A. Guanine
B. Cytosine
C. Thymine
D. Uracil

Q42. Telomerase action is more common in:
✅ A. Germ cells and cancer cells
B. RBCs
C. Muscle fibers
D. Platelets

Q43. Which sequence is typical of telomeres across many eukaryotes?
✅ A. G-rich repeats
B. TATA box
C. Palindromic repeats
D. Splice donor sites

Q44. Telomere overhang length is typically:
A. 2–4 bases
✅ B. 12–16 nucleotides
C. 50–100 bases
D. 1000 bp

Q45. Telomeric proteins TRF1 and TRF2 bind to:
✅ A. Double-stranded telomeric DNA
B. RNA template
C. Telomerase RNA
D. Origin of replication

Q46. The major difference between centromeric and telomeric DNA is:
✅ A. Telomeric DNA is conserved; centromeric is not
B. Centromere has no proteins
C. Telomeres don’t repeat
D. Telomerase acts on centromere

Q47. The kinetochore complex is absent in:
A. Centromere
✅ B. Telomere
C. Satellite DNA
D. Promoter

Q48. Telomeric DNA is associated with which nuclear structure?
✅ A. Nuclear lamina
B. Nucleolus
C. Ribosome
D. Centriole

Q49. Mutation in telomere regulatory proteins may lead to:
✅ A. Uncontrolled telomere elongation
B. Chromatin condensation
C. Transcription blockage
D. No effect

Q50. Which of the following is not a telomere-binding protein?
A. Rap1p
B. Taz1p
C. TRF
✅ D. CBF-III